Lecture 07: Electron transport/ATP production Respiration & Photophosphorylation - Biology

Lecture 07:  Electron transport/ATP production Respiration & Photophosphorylation - Biology

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Lecture 07: Electron transport/ATP production Respiration & Photophosphorylation

Electron transport chain

An electron transport chain (ETC) is a series of compounds that transfer electrons from electron donors to electron acceptors via redox reactions, and couples this electron transfer with the transfer of protons (H + ions) across a membrane. This creates an electrochemical proton gradient that drives the synthesis of adenosine triphosphate (ATP), a molecule that stores energy chemically in the form of highly strained bonds. The final acceptor of electrons in the electron transport chain is molecular oxygen.

Electron transport chains are used for extracting energy via redox reactions from sunlight in photosynthesis or, such as in the case of the oxidation of sugars, cellular respiration. In eukaryotes, an important electron transport chain is found in the inner mitochondrial membrane where it serves as the site of oxidative phosphorylation through the use of ATP synthase. It is also found in the thylakoid membrane of the chloroplast in photosynthetic eukaryotes. In bacteria, the electron transport chain is located in their cell membrane.

In chloroplasts, light drives the conversion of water to oxygen and NADP + to NADPH with transfer of H + ions across chloroplast membranes. In mitochondria, it is the conversion of oxygen to water, NADH to NAD + and succinate to fumarate that are required to generate the proton gradient.

Electron transport chains are major sites of premature electron leakage to oxygen, generating superoxide and potentially resulting in increased oxidative stress.

Reduction-Oxidation Reactions

In this class, most of the reduction/oxidation reactions (redox) that we discuss occur in metabolic pathways (connected sets of metabolic reactions). Here the cell breaks down the compounds it consumes into smaller parts and then reassembles them into larger macromolecules. Redox reactions also play critical roles in energy transfer, either from the environment or within the cell, in all known forms of life. For these reasons, it is important to develop at least an intuitive understanding and appreciation for redox reactions in biology.

Most students of biology will also study reduction and oxidation reactions in their chemistry courses these kinds of reactions are important well beyond biology. Regardless of the order in which students are introduced to this concept (chemistry first or biology first), most will find the topic presented in very different ways in chemistry and biology. That can be confusing.

Chemists often introduce the concepts of oxidation and reduction from the technically more correct and inclusive standpoint of oxidation states. See this link for more information: < ation_Numbers)>. Fortunately, there&rsquos no need to go into the details here (most of you will see that in chemistry at some point), just follow the argument for now. It might make things less confusing in both the long and short run. Anyhow, chemists will often ask students to apply a set of rules (see link above) to determine the oxidation states of individual atoms in a reaction. The chemistry formalism defines oxidation as an increase in oxidation state and reduction as a decrease in oxidation state.

All of this, of course, holds true in biology. However, biologists don&rsquot typically think of redox reactions in this way. Why? We suspect it&rsquos because most of the redox reactions encountered in biology involve a change in oxidation state that comes about because electrons are transferred between molecules. So, biologists typically define reduction as a gain of electrons and oxidation as a loss of electrons. The biological concept of redox is entirely consistent with the concept chemists use but it doesn&rsquot account for redox reactions that can happen without the transfer of electrons. The biologist&rsquos definition is therefore not as general as the chemist&rsquos, but it works for most cases encountered in biology.

This is a biology reading for a biology class. We, therefore, approach redox from the &ldquogain/loss of electrons&rdquo conceptualization that is commonly taught in biology classes. In our opinion, it&rsquos easier to use (no long list of rules to memorize and apply), more intuitive, and works for almost all cases we care about in undergraduate biology. So, if you had chemistry already and this topic seems a little different in biology, remember that at its core it&rsquos the same thing you learned about before. Biologists just adapted what you learned in chemistry to make more intuitive sense in biology. If you haven&rsquot learned about redox yet don&rsquot worry. If you can understand what we&rsquore trying to do here when you cover this concept in chemistry class you&rsquoll be a few steps ahead. You&rsquoll just need to generalize your thinking a bit instead of seeing the topic for the first time.

Let's start with some generic reactions

Transferring electrons between two compounds results in one of these compounds loosing an electron and one compound gaining an electron. For example, look at the figure below. If we use the energy story rubric to look at the overall reaction, we can compare the before and after characteristics of the reactants and products. What happens to the matter (stuff) before and after the reaction? Compound A starts as neutral and becomes positively charged. Compound B starts as neutral and becomes negatively charged. Because electrons are negatively charged, we can explain this reaction with the movement of an electron from Compound A to B. That is consistent with the changes in charge. Compound A loses an electron (becoming positively charged), and we say that A has become oxidized. For biologists, oxidation is associated with the loss of electron(s). B gains the electron (becoming negatively charged), and we say that B has become reduced. Reduction is associated with the gain of electrons. We also know, since a reaction occurred (something happened), that energy must have been transferred and/or reorganized in this process and we'll consider this shortly.

Figure 1.Generic redox reaction with half-reactions

To reiterate: When an electron(s) is lost, or a molecule is oxidized, the electron(s) must then pass to another molecule. We say that the molecule gaining the electron becomes reduced. Together these paired electron gain-loss reactions are known as an oxidation-reduction reaction (also called a redox reaction).

This idea of paired half-reactions is critical to the biological concept of redox. Electrons don&rsquot drop out of the universe for &ldquofree&rdquo to reduce a molecule or jump off a molecule into the ether. Donated electrons MUST come from a donor molecule and be transferred to some other acceptor molecule. For example in the figure above the electron the reduces molecule B in half-reaction 2 must come from a donor - it just doesn't appear from nowhere! Likewise, the electron that leaves A in half-reaction 1 above just "land" on another molecule - it doesn't just disappear from the universe.

Therefore, oxidation and reduction reactions must ALWAYS be paired. We&rsquoll examine this idea in more detail below when we discuss the idea of &ldquohalf-reactions&rdquo.

A tip to help you remember: The mnemonic LEO says GER (Lose Electrons = Oxidation and Gain Electrons = Reduction) can help you remember the biological definitions of oxidation and reduction.

Figure 2. A figure for the mnemonic "LEO the lion says GER." LEO: Loss of Electrons = Oxidation. GER: Gain of Electrons = Reduction

Attribution: Kamali Sripathi

&bull The vocabulary of redox can be confusing: Students studying redox chemistry can often become confused by the vocabulary used to describe the reactions. Terms like oxidation/oxidant and reduction/reductant look and sound very similar but mean distinctly different things. An electron donor is also sometimes called a reductant because it is the compound that causes the reduction (gain of electrons) of another compound (the oxidant). In other words, the reductant is donating it&rsquos electrons to the oxidant which is gaining those electrons. Conversely, the electron acceptor is called the oxidant because it is the compound that is causing the oxidation (loss of electrons) of the other compound. Again, this simply means the oxidant is gaining electrons from the reductant who is donating those electrons. Confused yet?

Yet another way to think about definitions is to remember that describing a compound as reduced/oxidized is describing the state that the compound itself is in, whereas labeling a compound as a reductant/oxidant describes how the compound can act, to either reduce or oxidize another compound. Keep in mind that the term reductant is also synonymous with reducing agent and oxidant is also synonymous with oxidizing agent. The chemists who developed this vocabulary need to be brought up on charges of "willful thickheadedness" at science trial and then be forced to explain to the rest of us why they needed to be so deliberately obtuse.

The confusing language of redox: quick summary

1. A compound can be described as &ldquoreduced&rdquo - term used to describe the compound's state

2. A compound can be a &ldquoreductant&rdquo - term used to describe a compound's capability (it can reduce something else). The synonymous term "reducing agent" can be used to describe the same capability (the term "agent" refers to the thing that can "do something" - in this case reduce another molecule).

3. A compound can be an &ldquooxidant&rdquo - term used to describe a compound's capability (it can oxidize something else). The synonymous term "oxidizing agent" can be used to describe the same capability (the term "agent" refers to the thing that can "do something" - in this case oxidize another molecule).

4. A compound can &ldquobecome reduced&rdquo or "become oxidized"- term used to describe the transition to a new state

Since all of these terms are used in biology, in General Biology we expect you to become familiar with this terminology. Try to learn it and use it as soon as possible - we will use the terms frequently and will not have the time to define terms each time.

Knowledge Check Quiz

The Half Reaction

Here we introduce the concept of the half reaction. We can think each half reaction as a description of what happens to one of the two molecules (i.e. the donor and the acceptor) involved in a "full" redox reaction. A "full" redox reaction requires two half reactions. We illustrate this below. In the example below, half reaction #1 depicts the molecule AH becoming losing two electrons and a proton and in the process becoming A + . This reaction depicts the oxidation of AH. Half reaction #2 depicts the molecule B + gaining two electrons and a proton to become BH. This reaction depicts the reduction of B + . Each of these two half reactions is conceptual and can't happen on their own. The electrons lost in half reaction #1 MUST go somewhere, they can't just disappear. Likewise, the electrons gained in half reaction #2 must come from something. They too just can't appear out of nowhere.

One can imagine that there might be different molecules that can serve as potential acceptors (the place for the electrons to go) for the electrons lost in half reaction #1. Likewise, there might be many potential reduced molecules that can serve as the electron donors (the source of electrons) for half reaction #2. In the example below, we show what happens (the reaction) when molecule AH is the donor of electrons for molecule B + . When we put the donor and acceptor half reactions together, we get a "full" redox reaction that can actually happen. In the figure below we call that reaction "Reaction #1". When this happens we call the two half reactions coupled.

Figure 3. Generic redox reaction where compound AH is being oxidized by compound B + . Each half reaction represents a single species or compound to either lose or gain electrons (and a subsequent proton as shown in the figure above). In half reaction #1 AH loses a proton and 2 electrons: in the second half reaction, B + gains 2 electrons and a proton. In this example HA is oxidized to A + while B + is reduced to BH.

Using this idea, we can theoretically couple and think about any two half reactions, one half reaction serving as the electron donor for the other half reaction that accepts the donated electrons. For instance, using the example above, we could consider coupling the reduction of B + that happens in half reaction 2 with another half reaction describing the oxidation of the molecule NADH. In that case, the NADH would be the electron donor for B + . Likewise you could couple the oxidation of AH that happens in half reaction #1 with a half reaction describing the the reduction of hypothetical molecule Z + . You can mix-and-match half reactions together as you please provided one half is describing the oxidation of a compound (it's donating electrons) and the reduction of another compound (it's accepting the donated electrons).

A note on how we write full reactions versus half reactions: In the example above, when we write Reaction #1 as an equation, the 2 electrons and the H + that are explicitly described in the underlying half reactions, are not explicitly included in the text of the full reaction. In the reaction above you must infer that an exchange of electrons happens. This can be observed by trying to balance charges between each reactant and it's corresponding product. Reactant AH becomes product A + . In this case, you can infer that some movement of electrons must have taken place. To balance the charges on this compound (make the sum of charges on each side of the equation equal) you need to add 2 electrons to the right side of the equation, one to account for the "+" charge on A + and a second to go with the H + that was also lost. The other reactant B + is converted to BH. It must therefore gain 2 electrons to balance charges, one for B + and a second for the additional H + that was added. Together this information leads you to conclude that the most likely thing to have happened is that two electrons were exchanged between AH and B + .

This will also be the case for most redox reactions in biology. Fortunately, in most cases, either the context of the reaction, the presence of chemical groups often engaged in redox (e.g. metal ions), or the presence of commonly used electron carriers (e.g. NAD + /NADH, FAD + /FADH2, ferredoxin, etc.) will alert you that the reaction is of class "redox". You will be expected to learn to recognize some of these common molecules.

Reduction Potential

By convention, we quantitatively characterize redox reactions using an measure called reduction potentials.The reduction potential attempts to quantitatively describe the &ldquoability&rdquo of a compound or molecule to gain or lose electrons. The specific value of the reduction potential is determined experimentally, but for the purpose of this course we assume that the reader will accept that the values in provided tables are reasonably correct. We can anthropomorphize the reduction potential by saying that it is related to the strength with which a compound can &ldquoattract&rdquo or &ldquopull&rdquo or &ldquocapture&rdquo electrons. Not surprisingly this is is related to but not identical to electronegativity.

What is this intrinsic property to attract electrons?

Different compounds, based on their structure and atomic composition have intrinsic and distinct attractions for electrons. This quality leads each molecule to have its own standard reduction potential or E0&rsquo. The reduction potential is a relative quantity (relative to some &ldquostandard&rdquo reaction). If a test compound has a stronger "attraction" to electrons than the standard (if the two competed, the test compound would "take" electrons from the standard compound), we say that the test compound has a positive reduction potential. The magnitude of the difference in E0&rsquo between any two compounds (including the standard) is proportional to how much more or less the compounds "want" electrons. The relative strength of the reduction potential is measured and reported in units of Volts (V)(sometimes written as electron volts or eV) or milliVolts (mV). The reference compound in most redox towers is H2.

Possible NB Discussion Point

Rephrase for yourself: How do you describe or think about the difference between the concept of electronegativity and red/ox potential?

Redox student misconception alert: The standard redox potential for a compound reports how strongly a substance wants to hold onto an electron in comparison to hydrogen. Since both redox potential and electronegativity are both discussed as measurements for how strongly something "wants" an electron, they are sometimes conflated or confused for one another. However, they are not. While the electronegativity of atoms in a molecule may influence its redox potential, it is not the only factor that does. You don't need to worry about how this works. For now, try to keep them as different and distinct ideas in your mind. The physical relationship between these two concepts is well beyond the scope of this general biology class.

The Redox Tower

All kinds of compounds can take part in redox reactions. Scientists have developed a graphical tool, the redox tower, to tabulate redox half reactions based on their E0 ' values. This tool can help predict the direction of electron flow between potential electron donors and acceptors and how much free energy change might be expected from a specific reaction. By convention, all half reactions in the table are written in the direction of reduction for each compound listed.

In the biology context, the electron tower usually ranks a variety of common compounds (their half reactions) from most negative E0 ' (compounds that readily get rid of electrons), to the most positive E0 ' (compounds most likely to accept electrons). The tower below lists the number of electrons that are transferred in each reaction. For example, the reduction of NAD + to NADH involves two electrons, written in the table as 2e - .

oxidized form

reduced form

n (electrons)

Eo´ (volts)

Ch.07- KEY Cellular respiration & Ferment In Focus-1

Catabolic Pathways are pathways that break down organic molecules, releasing stored energy.

Organic molecules possess potential energy as a result of the arrangement of electrons in the bonds between their atoms.

Fermentation is the partial breakdown of sugars without O 2 (anaerobic)

Aerobic respiration is the most efficient catabolic pathway. This occurs in the presence of O 2 which makes this an aerobic process.

The term cellular respiration includes both aerobic and anaerobic processes. However, it originated as a synonym for aerobic respiration because of the relationship to respiration and the act of breathing oxygen. Therefore, cellular respiration is often used to describe the aerobic process.

The basic breakdown of sugar can be represented through the chemical equation: C 6 H 1206 + 6 O 2 6 CO 2 + 6 H 2 O + NRG

***This reaction is an Exergonic reaction. G is negative in this reaction which means: Energy is released and this reaction is spontaneous.

Draw a graph that shows this reaction (show reactants, products, activation

  1. When cells break down glucose, the relocation of electrons releases energy stored in organic molecules, and this energy is used to synthesize ATP.

Oxidation / Reduction Reactions (OIL RIG)

In a redox reaction, the loss of electrons from one substance is called oxidation, and the addition of electrons is called reduction.

The electron donor is called the reducing agent while the electron acceptor is called the oxidizing agent.

An electron loses potential energy when it shifts from a less electronegative atom toward a more electronegative atom (energy is released,

Enzymes called dehydrogenases remove a pair of hydrogen atoms ( electrons and 2 protons) from the substrate, thereby oxidizing it.

The above molecule is a great intermediate because very little energy is lost when electrons (e-) are transferred from food to NAD+.

Electrons do not move directly from sugar to oxygen to harness energy to make ATP. The primary electron acceptor is NAD+ which delivers 2 electrons and 1 proton to the electron transport chain. FAD is another electron acceptor, but will deliver less energy to the electron transport chain.

Rocket fuel contains H 2 and O 2 and the energy used to power a rocket comes when the electrons from hydrogen “fall” closer to the more electronegative oxygen.

The basic “downhill” path that electrons follow as their energy is harnessed to produce ATP is: Food_ NADH or FADH 2 ETC O 2


Fill in the diagram below indicating the three stages of respiration. Indicate the molecule produced and method of production of ATP in each process.

The first two stages of cellular respiration, glycolysis and Krebs cycle are catabolic pathways that break down Glucose and other organic fuels.

Glycolysis which occurs in the cytosol begins the degradation process by breaking glucose into two molecules of pyruvate (pyruvic acid) GYCOLYSIS OVERVIEW (CAMPBELL)

The citric acid cycle (Krebs cycle) which takes place in the mitochondrial matrix of eukaryotic cells or the cytosol of prokaryotic cells, completes the breakdown of glucose by oxidizing pyruvate into carbon dioxide and water.

Through cellular respiration the energy currency (ATP) of the cell can be produced through two reactions: 27. Oxidative Phosphorylation which occurs in the electron transport chain with oxygen as the final electron acceptor. (accounts for 90 % of ATP production)

  1. The (net) energy result of this process is 2ATP plus 2NADH which will be used later by the electron transport chain to produce ATP if oxygen is present.

After pyruvate is oxidized, the Citric Acid Cycle (Krebs Cycle) completes the energy yielding oxidation of organic molecules CITRIC ACID CYCLE (CAMPBELL)

Glycolysis will release less than 25% of the energy stored in glucose while most of the energy will be stockpiled in the 2 molecules of pyruvate, which will be OXIDIZED/ REDUCED (CIRCLE ONE) if oxygen is present, or it will go through the process of fermentation if no oxygen is present.

Krebs Cycle: occurs in the mitochondrial matrix of eukaryotic cells.

The link stage between glycolysis and the Krebs cycle occurs when pyruvate is converted to actyl-Co A by the addition of coenzyme A.

Through this process: a. CO 2 is released as a waste.

b. NAD+ is reduced to produce NADH

Krebs cycle- occurs in the mitochondrial matrix

The one ATP molecule that is produced in each turn of the Krebs cycle is produced through Substrate level Phosphorylation.

3 molecules of NADH and 1 molecule of FADH 2 are formed in each turn of the cycle.

It takes 2 turns of the Krebs cycle to completely oxidize one molecule of glucose because of the 2 molecules of pyruvate made in glycolysis

Therefore, a total of 2 ATPs, 8 NADHs and 2 FADH 2 s are produced during the Krebs cycle including the conversion of pyruvate to Acetyl-CoA.

2 ATPs, 6 NADHs and 2 FADH 2 s are produced in Krebs ALONE.

The final electron acceptor is oxygen which is highly electronegative and becomes reduced to form water when it picks up a pair of hydrogen ions.

As electrons drop down the ETC energy is released and H+ is pumped against its concentration gradient from the matrix to the inner membrane space (pH in the inner membrane space is lower than in the matrix)

What is the difference between NADH and FADH 2 with regard to the ETC? NADH donates its electrons to the first electron acceptor in the ETC allowing the electrons to fall a greater distance and release maximum energy before it is accepted by O 2. Conversely, FADH 2 donates its electrons in the middle of the ETC. These electrons fall a shorter distance to O 2 and release less energy to drive chemiosmosis.

The electron transport chain makes 26-28 ATP directly

Chemiosmosis : The Energy-Coupling Mechanism

  1. All over the inner membrane of the mitochondrion or the prokaryotic plasma membrane are many copies of protein complex called ATP synthase that actually makes ATP from ADP and inorganic phosphate.

ATP synthase uses the energy of an existing ion gradient to power ATP synthesis.

The power source for the ATP synthase is a difference in the concentration of H+ ions on opposite sides of the inner mitochondrial membrane which were pumped during electron transport.

Chemiosmosis is the energy- coupling process by which a H+ ion gradient is used to create energy to produce ATP.

The H+ gradient is referred to as a proton motive force.


No Oxygen, No ATP, Not True.


Fermentation - is the process by which the anaerobic catabolism of nutrients can occur for an extended period of time.(it is an extension of glycolysis)

In order for the above process to occur, addtional, reactions are needed that regenerate NAD+ by transferring electrons to pyruvate.

The NAD+ can then be reused to oxidize the next glucose molecule by glycolysis.

Fermentation produces NO additional ATP molecules but allows more energy to be produced by the continued breakdown of glucose in glycolysis.

In alcohol fermentation, pyruvate is converted to ethanol (ethyl alcohol) and CO 2 is given off as a waste gas. This regenerates NAD+ which is needed for the continuation of glycolysis.

Many bacterial and yeast carry out alcoholic fermentation. Humans use these organisms for brewing, winemaking, and baking.

During lactic acid fermentation, pyruvate is reduced directly by NADH to form lactate, with NO release of CO 2. Human muscle cells use this process when Oxygen is scarce.

The lactate that accumulates was previously thought to cause muscle fatigue and pain, but recent research suggests that increased levels of potassium (K+) ions may be to blame. Excess lactate is carried away by the blood to the liver where it is gradually converted back to pyruvate.

Obligate anerobes are organisms that carry out only fermentation or anaerobic respiration and cannot survive in the presence of oxygen.

Other organisms can make enough ATP by using either fermentation or respiration are called facultative anaerobes.

Ancient prokaryotes probably used glycolysis to make ATP long before oxygen was present in the Earth’ atmosphere.

Glucose is not the only source of energy. Carbohydrates, proteins and fats can all be used for fuel. Label the following diagram.

The oxidation of fats will produce twice as much ATP as the same mass of carbohydrates.

Lab 07 - Cell. Respiration and Fermentation

Notification: If you have a disability that makes it difficult to complete this lab, please contact your instructor. Please provide your instructor a copy of the Memorandum of Accommodation (MOA) from NVCC Disability Support Services.


x Distinguish between anaerobic and aerobic respiration. x Measure the rate of oxygen consumption by living organisms during aerobic cellular respiration. x Calculate metabolic rate from the experimental data. x Measure the rate of yeast fermentation.


All life requires energy to survive. The energy of glucose is used as fuel by nearly all living organisms to stay alive, to maintain homeostasis, and run all kinds of life activities. The energy stored in chemical bonds of glucose and other food molecules needs to be released and converted into a diffusible, usable form ATP. When oxygen is present, most living organisms prefer to use the aerobic respiration pathway because they can make more energy (ATP) than when they use the anaerobic pathway. However, living organism can also partially break down glucose in the process of anaerobic respiration to produce 2 ATP molecules from each glucose molecule when oxygen is not available.

Aerobic respiration can produce up to 38 ATPs per molecule of glucose as compared to the 2 ATP that are generated in the anaerobic pathway. The three major parts of aerobic respiration are 1) Glycolysis, 2) Krebs Cycle (Citric Acid Cycle), and 3) Electron Transport Chain (ETC) and oxidative phosphorylation. The overall equation of cellular respiration is:

C 6 H 12 O 6 + 6 O 2 ĺ&amp2 2 + 6 H 2 O + 38 ATP

We can measure the rate of cellular respiration by measuring the consumption of the reactants (glucose or oxygen), or by measuring the rate of production of the end products (carbon dioxide or water) of this process. The easiest component to measure is oxygen consumption. If the living organism is placed in a closed system with the carbon dioxide removed as it is produced, then the consumption of oxygen can be determined using a respirometer. The respirometer consists of a glass container sealed with a rubber stopper outfitted with a pipette. A compound such as potassium hydroxide is placed in the respirometer to remove the carbon dioxide generated. As the living organism within consumes oxygen, water will displace the gas in the pipette and the volume of oxygen consumed can be measured over time to calculate the rate of cellular respiration and metabolic rate.

Anaerobic respiration is the incomplete break down of sugar. It produces lactic acid in animal cells and ethanol and carbon dioxide in yeast. Anaerobic respiration in yeast is also called fermentation, which is used to produce wine and bread, and more recently biofuels. Different

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sugars are used at different rates in yeast fermentation which can be measured by monitoring the production of carbon dioxide.


Follow all standard laboratory safety procedures. Be sure to wear gloves and use the forceps when working with potassium hydroxide.


Experiment 1. Cellular respiration

Ignore the use of worms and the use of glass beads in the video, we don’t use those in this lab activity.

Ignore the use of glass beads in the video, we don’t use those in this lab activity.

As oxygen is consumed by the organisms, water will start to enter the pipettes. See the image below for a depiction of the respirometer setup.

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Experiment 2. Yeast fermentation

  1. Watch the following video about setting up yeast fermentation tubes and measuring the height of the bubble at the top of the tubes from time 0:00 until 1:30 and from time 2:05 until 2:25.
  1. In the laboratory, you would obtain and label four small test tubes from 1 to 4. You would then fill them half full with the following solutions:

After putting the solutions into the test tubes, you would then fill all four tubes to the top with yeast suspension.

Using the results diagrammed below, fill in Table 2 in the Lab Worksheet

First, measure the height of the air bubble (in millimeters) in each tube at the Initial gas height and the Final gas height times and record your measurements in Table 2 in the lab worksheet.

Then, determine the amount of carbon dioxide produced by yeast fermentation in each of the tubes by subtracting the height of the air bubble at the Initial gas height time from the height of the air bubble at the Final gas height time. Enter your results in Table 2 in the Lab Worksheet.

For example, if the height of the air bubble at the Final gas height time is 25 mm and the height of the air bubble at the Initial gas height time is 5 mm, then the change in height, which corresponds to the amount of CO 2 produced, is 25 mm

upload their completed document as a DOC or PDF file in Canvas

upload their completed document as a DOC or PDF file in Canvas

Table 2. Yeast fermentation

Tube Sugar Initial gas height (mm)

Final gas height (mm)

Change in height (mm)

What is the independent variable for the yeast fermentation experiment?

What is the dependent variable for the yeast fermentation experiment?

Which sugar resulted in the fasted production of CO2 gas? What is your explanation for this observation?

24mm 17mm 7mm

22mm 14mm 8mm

Lecture 07: Electron transport/ATP production Respiration & Photophosphorylation - Biology

INTRODUCTION Under appropriate conditions, pyruvate can be further oxidized. One of the most studied oxidation reactions involving pyruvate is a two part reaction involving NAD + and molecule called co-enzyme A (CoA). This reaction oxidizes pyruvate, leads to a loss of one carbon via decarboxylation, and creates a new molecule called acetyl-CoA. The resulting acetyl-CoA can enter several pathways for the biosynthesis of larger molecules or it can be routed to another pathway of central metabolism called the citric acid cycle. Here the remaining two carbons can either be further oxidized or serve again as precursors for the construction of various other molecules. We discuss these scenarios below.

The different fates of Pyruvate

Module 5.3 left off with the end-products of glycolysis: 2 pyruvate molecules, 2 ATPs and 2 NADH molecules. This module and module 5.5 will explore what the cell may now do with the pyruvate, ATP and NADH that were generated. In module 5.5 we will see how pyruvate is the primary starting substrate for fermentation reactions, reactions that allow cells to regenerate NAD + from NADH, to allow for the continued oxidation of glucose and the uninterrupted continuation of glycolysis. In this module, we will explore the continued and complete oxidation of pyruvate all the way to CO2.

The fates of ATP and NADH In general, ATP can be used for or coupled to a variety of cellular functions including biosynthesis, transport, replication etc. We will see many such examples.

What to do with the NADH however, depends on the conditions under which the cell is growing. In some cases, the cell will opt to rapidly recycle NADH back into to NAD + . This occurs through a process called fermentation in which the electrons initially taken from the glucose derivatives are returned to more downstream products via another redox transfer (described in more detail in module 5.5). Alternatively, NADH can be recycled back into NAD + by donating electrons to something known as an electron transport chain (this is covered in module 5.6).

  • Pyruvate can be used as a terminal electron acceptor (either directly or indirectly) in fermentation reactions, and is discussed in Module 5.5.
  • Pyruvate could be secreted from the cell as a waste product.
  • Pyruvate could be further oxidized to extract more free energy from this fuel.

The further oxidation of pyruvate In respiring bacteria and archaea, the pyruvate is further oxidized in the cytoplasm. In aerobically respiring eukaryotic cells, the pyruvate molecules produced at the end of glycolysis are transported into mitochondria, which are sites of cellular respiration and house oxygen consuming electron transport chains (ETC in module 5.6). Organisms from all three domains of life share similar mechanisms to further oxidize the pyruvate to CO2. First pyruvate is decarboxylated and covalently linked to co-enzyme A via a thioester linkage to form the molecule known as acetyl-CoA . While acetyl-CoA can feed into multiple other biochemical pathways we now consider its role in feeding the circular pathway known as the Tricarboxylic Acid Cycle , also referred to as the TCA cycle , the Citric Acid Cycle or the Krebs Cycle . This process is detailed below.

Conversion of Pyruvate into Acetyl-CoA

The conversion of pyruvate into acetyl-CoA In a multistep reaction catalyzed by the enzyme pyruvate dehydrogenase, pyruvate is oxidized by NAD + , decarboxylated, and covalently linked to a molecule of co-enzyme A via a thioester bond. Remember: there are two pyruvate molecules produced at the end of glycolysis for every molecule of glucose metabolized thus, two of the six carbons will have been removed at the end of both steps. The release of the carbon dioxide is important here, this reaction often results in a loss of mass from the cell as the CO2 will diffuse or be transported out of the cell and become a waste product. In addition, NAD + is reduced to NADH during this process per molecule of pyruvate oxidized.

Upon entering the mitochondrial matrix, a multi-enzyme complex converts pyruvate into acetyl CoA. In the process, carbon dioxide is released and one molecule of NADH is formed.

In the presence of a suitable terminal electron acceptor, acetyl CoA delivers (exchanges a bond) its acetyl group to a four-carbon molecule, oxaloacetate, to form citrate (designated the first compound in the cycle). This cycle is called by different names: the citric acid cycle (for the first intermediate formed—citric acid, or citrate), the TCA cycle (since citric acid or citrate and isocitrate are tricarboxylic acids), and the Krebs cycle , after Hans Krebs, who first identified the steps in the pathway in the 1930s in pigeon flight muscles.

The Tricarboxcylic Acid (TCA) Cycle also called the Krebs cycle

In bacteria and archaea reactions in the citric acid cycle typically happen in the cytosol. In eukaryotes, the citric acid cycle takes place in the matrix of mitochondria. Almost all (but not all) of the enzymes of the citric acid cycle are water soluble (not in the membrane), with the single exception of the enzyme succinate dehydrogenase, which is embedded in the inner membrane of the mitochondrion (in eukaryotes). Unlike glycolysis, the citric acid cycle is a closed loop: the last part of the pathway regenerates the compound used in the first step. The eight steps of the cycle are a series of redox, dehydration, hydration, and decarboxylation reactions that produce two carbon dioxide molecules, one ATP, and reduced forms of NADH and FADH2.

In the citric acid cycle, the acetyl group from acetyl CoA is attached to a four-carbon oxaloacetate molecule to form a six-carbon citrate molecule. Through a series of steps, citrate is oxidized, releasing two carbon dioxide molecules for each acetyl group fed into the cycle. In the process, three NAD + molecules are reduced to NADH, one FAD molecule is reduced to FADH2, and one ATP or GTP (depending on the cell type) is produced (by substrate-level phosphorylation). Because the final product of the citric acid cycle is also the first reactant, the cycle runs continuously in the presence of sufficient reactants. (credit: modification of work by “Yikrazuul”/Wikimedia Commons)

Steps in the Citric Acid Cycle

Step 1. The first step of the cycle is a condensation reaction involving the two-carbon acetyl group of acetyl-CoA with one four-carbon molecule of oxaloacetate. The products of this reaction are the six-carbon molecule citrate and free co-enzyme A. This step is considered irreversible because it is so highly exergonic. Moreover, the rate of this reaction is controlled through negative feedback by ATP. If ATP levels increase, the rate of this reaction decreases. If ATP is in short supply, the rate increases. If not already, the reason will become evident shortly.

Step 2. In step two, citrate loses one water molecule and gains another as citrate is converted into its isomer, isocitrate.

Step 3. In step three, isocitrate is oxidized by NAD + and decarboxylated. Keep track of the carbons! This carbon now more than likely leaves the cell as waste and is no longer available for building new biomolecules. The oxidation of isocitrate therefore produces a five-carbon molecule, α-ketoglutarate, a molecule of CO2 and reduced NADH. This step is also regulated by negative feedback from ATP and NADH, and a positive effect from ADP.

Step 4. Step 4 is catalyzed by the enzyme succinate dehydrogenase. Here, α-ketoglutarate is further oxidized by NAD + . This oxidation again leads to a decarboxylation and thus the loss of another carbon as waste. So far two carbons have come into the cycle from acetyl-CoA and two have left as CO2. At this stage, There is no net gain of carbons assimilated from the glucose molecules that are oxidized to this stage of metabolism. Unlike the previous step however succinate dehydrogenase - like pyruvate dehydrogenase before it - couples the free energy of the exergonic redox and decarboxylation reaction to drive the formation of a thioester bond between the substrate co-enzyme A and succinate (what is left after the decarboxylation). Succinate dehydrogenase is regulated by feedback inhibition of ATP, succinyl-CoA, and NADH.

Step 5. In step five, There is a substrate level phosphorylation event, where inorganic phosphate (Pi) is added to GDP or ADP to form GTP (an ATP equivalent for our purposes)or ATP. The energy that drives this substrate level phosphorylation event comes from the hydrolysis of the CoA molecule from succinyl

CoA to form succinate. Why is either GTP or ATP produced? In animal cells there are two isoenzymes (different forms of an enzyme that carries out the same reaction), for this step, depending upon the type of animal tissue in which they are found. One form is found in tissues that use large amounts of ATP, such as heart and skeletal muscle. This form produces ATP. The second form of the enzyme is found in tissues that have a high number of anabolic pathways, such as liver. This form produces GTP. GTP is energetically equivalent to ATP however, its use is more restricted. In particular, the process of protein synthesis primarily uses GTP. Most bacterial systems produce GTP in this reaction.

Step 6. Step six is another redox reactions in which succinate is oxidized by FAD + into fumarate. Two hydrogen atoms are transferred to FAD + , producing FADH2. The difference in reduction potential between the fumarate/succinate and NAD + /NADH half reactions is insufficient to make NAD + a suitable reagent for oxidizing succinate with NAD + under cellular conditions. However, the difference in reduction potential with the FAD + /FADH2 half reaction is adequate to oxidize succinate and reduce FAD + . Unlike NAD + , FAD + remains attached to the enzyme and transfers electrons to the electron transport chain (Module 5.6) directly. This process is made possible by the localization of the enzyme catalyzing this step inside the inner membrane of the mitochondrion or plasma membrane (depending on whether the organism in question is eukaryotic or not).

Step 7. Water is added to fumarate during step seven, and malate is produced. The last step in the citric acid cycle regenerates oxaloacetate by oxidizing malate with NAD + . Another molecule of NADH is produced in the process.

Summary Note that this process completely oxidizes 1 molecule of pyruvate, a 3 carbon organic acid, to 3 molecules of CO2. During this process, 4 molecules of NADH, 1 molecule of FADH2, and 1 molecule of GTP (or ATP) are produced. For respiring organisms this is a significant source of energy, since each molecule of NADH and FAD2 can feed directly into the electron transport chain, and as we will soon see, the subsequent redox reactions will indirectly energetically drive the synthesis of additional ATP. This suggests that the TCA cycle is primarily an energy generating mechanism evolved to extract or convert as much potential energy form the original energy source to a form cells can use, ATP (or the equivalent) or an energized membrane. However, - and let us not forget - the other important outcome of evolving this pathway is the ability to produce several precursor or substrate molecules necessary for various catabolic reactions (this pathway provides some of the early building blocks to make bigger molecules). As we will discuss below, there is a strong link between carbon metabolism and energy metabolism.

Click through each step of the citric acid cycle here.

Energy Stories

Work on building some energy stories There are a few interesting reactions that involve large transfers of energy and rearrangements of matter. Pick a few. Rewrite a reaction in your notes, and practice constructing an energy story. You now have the tools to discuss the energy redistribution in the context of broad ideas and terms like exergonic and endergonic. You also have the ability to begin discussing mechanism (how these reactions happen) by invoking enzyme catalysts. See your instructor and/or TA and check with you classmates to self-test on how you're doing.

Connections to Carbon Flow

One hypothesis that we have started exploring in this reading and in class is the idea that "central metabolism" evolved as a means of generating carbon precursors for catabolic reactions. Our hypothesis also states that as cells evolved, these reactions became linked into pathways: glycolysis and the TCA cycle, as a means to maximize their effectiveness for the cell. A side benefit to this evolving metabolic pathway was the generation of NADH from the complete oxidation of glucose - we saw the beginning of this idea when we discussed fermentation. We have already discussed how glycolysis not only provides ATP from substrate level phosphorylation, but also yields a net of 2 NADH molecules and 6 essential precursores: glucose-6-P, fructose-6-P, trios-P, 3-phosphoglycerate, phosphoenolphyruvate, and of course pyruvate. While ATP can be used by the cell directly as an energy source, NADH posses a problem and must be recycled back into NAD + , to keep the cycle in balance. As we see in detail in module 5.5, the most ancient way cells deal with this poblem is to use fermentation reactions to regenerate NAD + .

During the process of pyruvate oxidation via the TCA cycle 4 additional essential precursors are formed: acetyle

CoA, alpha-ketoglutarate, oxaloacetate, and succinyl

CoA. Three molecules of CO2 are lost and this represents a net loss of mass for the cell. These precursors, however, are substrates for a variety of catabolic reactions including the production of amino acids, fatty acids, and various co-factors, such as heme. This means that the rate of reaction through the TCA cycle will be sensitive to the concentrations of each metabolic intermediate (more on the thermodynamics in class). A metabolic intermediate is a compound that is produced by one reaction (a product) and then acts as a substrate for the next reaction. This also means that metabolic intermediates, in particular the 4 essential precursors, can be removed at any time for catabolic reactions, if there is a demand.

Not all cells have a functional TCA cycle Since all cells require the ability of make these precursor molecules, one might expect that all organisms would have a fully functional TCA cycle. In fact, the cells of many organisms DO NOT have a the enzymes to form a complete cycle - all cells, however, DO have the capability of making the 4 TCA cycle precursors noted in the previous paragraph. How can the cells make precursors and not have a full cycle? Remember that most of these reactions are freely reversible, so, if NAD + is required to for the oxidation of pyruvate or acetyl

CoA, then the reverse reactions would require NADH. This process is often referred to as the reductive TCA cycle. To drive these reactions in reverse (with respect to the direction discussed above) requires energy, in this case carried by ATP and NADH. If you get ATP and NADH driving a pathway one direction, it stands to reason that driving it in reverse will require ATP and NADH as "inputs".

Additional Links

Here are some additional links to videos and pages that you may find useful.

Additional Questions:

What is the primary difference between a circular pathway and a linear pathway?

In a circular pathway, the final product of the reaction is also the initial reactant. The pathway is self-perpetuating, as long as any of the intermediates of the pathway are supplied. Circular pathways are able to accommodate multiple entry and exit points, thus being particularly well suited for amphibolic pathways. In a linear pathway, one trip through the pathway completes the pathway, and a second trip would be an independent event.

MODULE 05.6 Oxidative Phosphorylation and the Electron Transport Chain

INTRODUCTION The electron transport chain (ETC) is the portion of respiration that uses an external electron acceptor as the final/terminal acceptor for the electrons that were removed from the intermediate compounds in glucose catabolism. In eukaryotic cells the ETC is composed of four large, multiprotein complexes embedded in the inner mitochondrial membrane and two small diffusible electron carriers shuttling electrons between them. The electrons are passed from enzyme to enzyme through a series of redox reactions. These reactions are couple the exergonic redox transfers to the endergonic transport of hydrogen ions across the membrane. This process contributes to the creation of a transmembrane electrochemical gradient. The electrons passing through the ETC gradually lose potential energy up until the point they are deposited on the terminal electron acceptor. The free energy difference of this multistep redox process is

-60 kcal/mol when NADH donates electrons or 45 kcal/mol when FADH2 donates, for organisms using oxygen as the final electron acceptor.

Introduction to Red/Ox, oxidative phosphorylation and Electron Transport Chains In modules 5.1, we discussed the general concept of Red/Ox reactions in biology and introduced the Electron Tower, a tool to help you understand Red/Ox chemistry and to estimate the direction and magnitude of potential energy differences for various Red/Ox couples. In modules 5.3 and 5.4 substrate level phosphorylation and fermentation were discussed and we saw how exergonic Red/Ox reactions could be directly coupled by enzymes to the endergonic synthesis of ATP. These processes are hypothesized to be one of the oldest forms of energy production used by cells. In this section we discuss the next evolutionary advancement in cellular energy metabolism, oxidative phosphorylation. First and foremost, oxidative phosphorylation does not imply the use of oxygen, it can, but it does not have to use oxygen. It is called oxidative phosphorylation because it relies on Red/Ox reactions to generate a electrochemical transmembrane potential that can then be used by the cell to do work.

A quick summary of Electron Transport Chains The ETC begins with the addition of electrons, donated from NADH, FADH2 or other reduced compounds. These electrons move through a series of electron transporters, enzymes that are embedded in a membrane, or carriers that undergo Red/Ox reactions. The free energy transferred from these exergonic Red/Ox reactions is coupled to the endergonic movement of protons across a membrane. This unequal accumulation of protons on either side of the membrane "polarizes" or "charges" the membrane, with a net positive (protons) on one side of the membrane and a negative charge on the other side of the membrane. The separation of charge creates an electrical potential . In addition, the accumulation of protons also causes a pH gradient known as a chemical potential across the membrane. Together these two gradients (electrical and chemical) are called an electro-chemical gradient .

Review: The Electron Tower

Since Red/Ox chemistry is so central to the topic we begin with a quick review of the table of reduction potential - sometimes called the "redox tower". As we discussed in Module 5.1, all kinds of compounds can participate in biological Red/Ox reactions. Making sense of all of this information and ranking potential Red/Ox pairs can be confusing. A tool has been developed to rate Red/Ox half reactions based on their reduction potentials or E0 ' values. Whether a particular compound can act as an electron donor (reductant) or electron acceptor (oxidant) depends on what other compound it is interacting with. The redox tower ranks a variety of common compounds (their half reactions) from most negative E0 ' , compounds that readily get rid of electrons, to the most positive E0 ' , compounds most likely to accept electrons. The tower organizes these half reactions based on the ability of electrons to accept electrons. In addition, in many redox towers each half reaction is written by convention with the oxidized form on the left followed by the reduced form to its right. The two forms may be either separated by a slash, for example the half reaction for the reduction of NAD + to NADH is written: NAD + /NADH + 2e - , or by separate columns. An electron tower is shown in figure 1 below.

Common Red/ox tower

Review Red/Ox Tower video from Module 5.1 For a short video on how to use the redox tower in red/ox problems click here. This video was made by Dr. Easlon for Bis2A students.

Using the Red/Ox Tower: A tool to help understand electron transport chains

By convention the tower half reactions are written with the oxidized form of the compound on the left and the reduced form on the right. Notice that compounds such as glucose and hydrogen gas are excellent electron donors and have very low reduction potentials E0 ' . Compounds, such as oxygen and nitrite, whose half reactions have relatively high positive reduction potentials (E0 ' ) generally make good electron acceptors are found at the opposite end of the table.

Menaquinone: an example Let's look at menaquinoneox/red. This compound sits in the middle of the redox tower with an half-reaction E0 ' value of -0.074 eV. Menaquinoneox can spontaneously (ΔG<0) accept electrons from reduced forms of compounds with lower half-reaction E0 ' . Such transfers form menaquinonered and the oxidized form of the original electron donor. In the table above, examples of compounds that could act as electron donors to menaquinone include FADH2, an E0 ' value of -0.22, or NADH, with an E0 ' value of -0.32 eV. Remember the reduced forms are on the right hand side of the red/ox pair.

Once menaquinone has been reduced, it can now spontaneously (ΔG<0) donate electrons to any compound with a higher half-reaction E0 ' value. Possible electron acceptors include cytochrome box with an E0 ' value of 0.035 eV or ubiquinoneox with an E0 ' of 0.11 eV. Remember that the oxidized forms lie on the left side of the half reaction.

The Electron Transport Chain

The electron transport chain , or ETC , is composed of a group of protein complexes in and around a membrane that help couple to energetically couple a series of exergonic/spontaneous red/ox reactions to the endergonic pumping of protons across the membrane to generate a an electro-chemical gradient. This electrochemical gradient creates a free energy potential that is termed a proton motive force whose energetically "downhill" exergonic transfer can later be later coupled to a variety of cellular processes.

  1. Electrons enter the ETC from a high energy electron donor, such as NADH or FADH2, which are generated during a variety of catabolic reactions like and including those associated glucose oxidation (review modules 5.3-5.5). Depending on the complexity (number and types of electron carriers) of the ETC being used by an organism, electrons can enter at a variety of places in the electron transport chain - this depends upon the respective reduction potentials of the proposed electron donors and acceptors.
  2. After the first redox reaction, the initial electron donor will become oxidized and the electron acceptor will become reduced. The difference in redox potential between the electron acceptor an donor is related to ΔG by the relationship ΔG = -nFΔE, where n = the number of electrons transferred and F = Faraday's constant. The larger a positive ΔE the more exergonic a reaction.
  3. If sufficient energy transferred during an exergonic redox step the electron carrier may couple this negative change in free energy to the endergonic process of transporting a proton from one side of the membrane to the other.
  4. After multiple redox transfers, the electron is delivered to a molecule known as the terminal electron acceptor. In the case of humans and plants, this is oxygen. However, there are many, many, many, other possible electron acceptors, see below.

What are the complexes of the ETC? ETCs are made up of a series (at least one) of membrane associated red/ox proteins or (some are integral) protein complexes (complex = more than one protein arranged in a quaternary structure) that move electrons from a donor source, such as NADH, to a final terminal electron acceptor, such as oxygen - this donor/terminal acceptor pair is the primary one used in human mitochondria. Each electron transfer in the ETC requires a reduced substrate as an electron donor and an oxidized substrate as the electron acceptor. In most cases the electron acceptor is a member of the enzyme complex. Once the complex is reduced, the complex can serve as an electron donor for the next reaction.

How do ETC complexes transfer electrons? As previously mentioned the ETC is composed of a series of protein complexes that undergo a series of linked red/ox reactions. These complexes are in fact multiprotein enzyme complexes referred to as oxidoreductases or simply reductases . The one exception to this naming convention is the terminal complex in aerobic respiration that uses molecular oxygen as the terminal electron acceptor. That enzyme complex is referred to as an oxidase . Red/Ox reactions in these complexes are typically carried out by a non-protein moiety called a prosthetic group . This is true for all of the electron carriers with the exception of quinones, which are a class of lipids that can directly be reduced or oxidized by the oxidoreductases. In this case, both the Quinonered and the Quinoneox is soluble within the membrane and can move from complex to complex. The prosthetic groups are directly involved in the red/ox reactions being catalyzed by their associated oxidoreductases. In general these prosthetic groups can be divided into two general types: those that carry both electrons and protons and those that only carry electrons.

  • Flavoproteins ( Fp ), these proteins contain an organic prosthetic group called a flavin , which is the actual moiety that undergoes the oxidation/reduction reaction. FADH2 is an example of a Fp.
  • Quinones , are a family of lipids which means they are soluble within the membrane.
  • It should also be noted that NADH and NADPH are considered electron (2e-) and proton (2 H + ) carriers.
  • Cytochromes are proteins that contain a heme prosthetic group. The Heme is capable of carrying a single electron.
  • Iron-Sulfur proteins contain a non-heme iron-sulfur clusters that can carry an electron. The prosthetic group is often abbreviated as Fe-S

Aerobic versus Anaerobic respiration In the world we live in, most of the organisms we interact with breath air, which is approximately 20% oxygen. Oxygen is our terminal electron acceptor . We call this process respiration, specifically aerobic respiration, we breath in oxygen, our cells take it up and transport it into the mitochondria where it is used as the final acceptor of electrons from our electron transport chains. That is aerobic respiration : the process of using oxygen as a terminal electron acceptor in an electron transport chain.

While most of the organisms we interact with use oxygen as the terminal electron acceptor, this process of respiration evolved at time when oxygen was not a major component of the atmosphere. Respiration or oxidative phosphorylation does not require oxygen at all it simply requires a compound with a high reduction potential to act as a terminal electron acceptor accept electrons from one of the complexes within the ETC. Many organisms can use a variety of compounds including nitrate (NO3 - ), nitrite (NO2 - ), even iron (Fe +++ ) as terminal electron acceptors. When oxygen is NOT the terminal electron acceptor, the process is referred to as anaerobic respiration . The ability of an organism to vary its terminal electron acceptor provides metabolic flexibility and can ensure better survival if any given terminal acceptor is in limited supply. Think about this, in the absence of oxygen we die but an organism that can use a different terminal electron acceptor can survive.

A generic example of a simple, 2 complex ETC Figure 2 shows a generic electron transport chain, composed of two integral membrane complexes Complex Iox and Complex IIox. A reduced electron donor, designated DH (such as NADH or FADH2) reduces Complex 1ox giving rise to the oxidized form D (such as NAD or FAD). Simultaneously, a prosthetic group within complex I is now reduced (accepts the electrons). In this example the redox reaction is exergonic and the free energy difference is coupled by the enzymes in Complex I to the endergonic translocation of a proton from one side of the membrane to the other. The net result is that one surface of the membrane becomes more negatively charged, due to an excess of hydroxyl ions (OH - ) and the other side becomes positively charged due to an increase in protons on the other side. Complex Ired can now reduce the prosthetic group in Complex IIred while simultaneously oxidizing Complex Ired. Electrons pass from Complex I to Complex II via thermodynamically spontaneous red/ox reactions, regenerating Complex Iox which can repeat the previous process. Complex IIred reduces A, the terminal electron acceptor to regenerate Complex IIox and create the reduced form of the terminal electron acceptor. In this case, Complex II can also translocate a proton during the process. If A is molecular oxygen, water (AH) will be produced. This reaction would then be considered a model of an aerobic ETC. However, if A is nitrate, NO3 - then Nitrite, NO2 - is produced (AH) and this would be an example of an anaerobic ETC.

Generic 2 complex electron transport chain. In the figure, DH is the electron donor (donor reduced) and D is the donor oxidized. A is the oxidized terminal electron acceptor and AH is the final product, the reduced form of the acceptor. As DH is oxidized to D, protons are translocated across the membrane, leaving an excess of hydroxyl ions (negatively charged) on one side of the membrane and protons (positively charged) on the other side of the membrane. The same reaction occurs in Complex II as the terminal electron acceptor is reduced to AH.

Based on Figure 2 above and using the electron tower in Figure 1, what is the difference in the electrical potential if (A) DH is NADH and A is O2 and (B) DH is NADH and A is NO3 - . Which pairs (A or B) provide the most amount of usable energy?

Detailed look at aerobic respiration The eukaryotic mitochondria has evolved a very efficient ETC. There are four complexes composed of proteins, labeled I through IV in [link], and the aggregation of these four complexes, together with associated mobile, accessory electron carriers, is called the electron transport chain. The electron transport chain is present in multiple copies in the inner mitochondrial membrane of eukaryotes and the plasma membrane of bacteria and arechaea.

The electron transport chain is a series of electron transporters embedded in the inner mitochondrial membrane that shuttles electrons from NADH and FADH2 to molecular oxygen. In the process, protons are pumped from the mitochondrial matrix to the intermembrane space, and oxygen is reduced to form water.

Complex I

To start, two electrons are carried to the first complex aboard NADH. This complex, labeled I, is composed of flavin mononucleotide (FMN) and an iron-sulfur (Fe-S)-containing protein. FMN, which is derived from vitamin B2, also called riboflavin, is one of several prosthetic groups or co-factors in the electron transport chain. A prosthetic group is a non-protein molecule required for the activity of a protein. Prosthetic groups are organic or inorganic, non-peptide molecules bound to a protein that facilitate its function prosthetic groups include co-enzymes, which are the prosthetic groups of enzymes. The enzyme in complex I is NADH dehydrogenase and is a very large protein, containing 45 amino acid chains. Complex I can pump four hydrogen ions across the membrane from the matrix into the intermembrane space, and it is in this way that the hydrogen ion gradient is established and maintained between the two compartments separated by the inner mitochondrial membrane.

Q and Complex II

Complex II directly receives FADH2, which does not pass through complex I. The compound connecting the first and second complexes to the third is ubiquinone (Q). The Q molecule is lipid soluble and freely moves through the hydrophobic core of the membrane. Once it is reduced, (QH2), ubiquinone delivers its electrons to the next complex in the electron transport chain. Q receives the electrons derived from NADH from complex I and the electrons derived from FADH2 from complex II, including succinate dehydrogenase. This enzyme and FADH2 form a small complex that delivers electrons directly to the electron transport chain, bypassing the first complex. Since these electrons bypass and thus do not energize the proton pump in the first complex, fewer ATP molecules are made from the FADH2 electrons. As we will see in the following section, the number of ATP molecules ultimately obtained is directly proportional to the number of protons pumped across the inner mitochondrial membrane.

Complex III

The third complex is composed of cytochrome b, another Fe-S protein, Rieske center (2Fe-2S center), and cytochrome c proteins this complex is also called cytochrome oxidoreductase. Cytochrome proteins have a prosthetic group of heme. The heme molecule is similar to the heme in hemoglobin, but it carries electrons, not oxygen. As a result, the iron ion at its core is reduced and oxidized as it passes the electrons, fluctuating between different oxidation states: Fe ++ (reduced) and Fe +++ (oxidized). The heme molecules in the cytochromes have slightly different characteristics due to the effects of the different proteins binding them, giving slightly different characteristics to each complex. Complex III pumps protons through the membrane and passes its electrons to cytochrome c for transport to the fourth complex of proteins and enzymes (cytochrome c is the acceptor of electrons from Q however, whereas Q carries pairs of electrons, cytochrome c can accept only one at a time).

Complex IV

The fourth complex is composed of cytochrome proteins c, a, and a3. This complex contains two heme groups (one in each of the two cytochromes, a, and a3) and three copper ions (a pair of CuA and one CuB in cytochrome a3). The cytochromes hold an oxygen molecule very tightly between the iron and copper ions until the oxygen is completely reduced. The reduced oxygen then picks up two hydrogen ions from the surrounding medium to make water (H2O). The removal of the hydrogen ions from the system contributes to the ion gradient used in the process of chemiosmosis.


In chemiosmosis, the free energy from the series of redox reactions just described is used to pump hydrogen ions (protons) across the membrane. The uneven distribution of H + ions across the membrane establishes both concentration and electrical gradients (thus, an electrochemical gradient), owing to the hydrogen ions’ positive charge and their aggregation on one side of the membrane.

If the membrane were open to diffusion by the hydrogen ions, the ions would tend to diffuse back across into the matrix, driven by their electrochemical gradient. Many ions cannot diffuse through the nonpolar regions of phospholipid membranes without the aid of ion channels. Similarly, hydrogen ions in the matrix space can only pass through the inner mitochondrial membrane through an integral membrane protein called ATP synthase ([link]). This complex protein acts as a tiny generator, turned by transfer of energy mediated by protons moving down their electrochemical gradient. The movement of this molecular machine (enzyme) serves to lower the activation energy of reaction and couples the exergonic transfer of energy associated with the movement of protons down their electrochemical gradient to the endergonic addition of a phosphate to ADP, forming ATP.

ATP synthase is a complex, molecular machine that uses a proton (H + ) gradient to form ATP from ADP and inorganic phosphate (Pi). (Credit: modification of work by Klaus Hoffmeier)

Chemiosmosis ([link]) is used to generate 90 percent of the ATP made during aerobic glucose catabolism it is also the method used in the light reactions of photosynthesis to harness the energy of sunlight in the process of photophosphorylation. Recall that the production of ATP using the process of chemiosmosis in mitochondria is called oxidative phosphorylation. The overall result of these reactions is the production of ATP from the energy of the electrons removed from hydrogen atoms. These atoms were originally part of a glucose molecule. At the end of the pathway, the electrons are used to reduce an oxygen molecule to oxygen ions. The extra electrons on the oxygen attract hydrogen ions (protons) from the surrounding medium, and water is formed.

In oxidative phosphorylation, the pH gradient formed by the electron transport chain is used by ATP synthase to form ATP in a Gram - bacteria.

A Hypothesis as to how ETC may have evolved

A proposed link between SLP/Fermentation and the evolution of ETCs When we last discussed energy metabolism, it was in context of substrate level phosphorylation (SLP) and fermentation reactions. One of the questions in the Discussion points was what would be the consequences of SLP, both short-term and long-term to the environment? We discussed how cells would need to co-evolve mechanisms to remove protons from the cytosol (interior of the cell), which lead to the evolution of the F0F1ATPase, a multi-subunit enzyme that translocates protons from the inside of the cell to the outside of the cell by hydrolyzing ATP as shown in figure 6 below. This arrangement works as long as small reduced organic molecules are freely available, making SLP and fermentation advantageous. As these biological process continue, the small reduced organic molecules begin to be used up and their concentration decreases, putting a demand on cells to be more efficient. One source of potential "ATP waste" is in the removal of protons from the cell's cytosol, organisms that could find other mechanisms could have a selective advantage. Such selective pressure could have led to the first membrane-bound proteins that could use Red/Ox reactions as their energy source, as depicted in figuire 7 . In other words use the energy from a Red/Ox reaction to move protons. Such enzymes and enzyme complexes exist today in the form of the electron transport complexes, like Complex I, the NADH dehydrogenase.

Proposed evolution of an ATP dependent proton translocator As small reduced organic molecules become limiting organisms that can find alternative mechanisms to remove protons from the cytosol may have had and advantage. The evolution of a proton translocator that uses the energy in a Red/Ox reaction could substitute for the ATAase.

Continuing with this line of logic, there are organisms that can now use Red/Ox reactions to translocate protons across the membrane, instead of an ATP driven proton pump. With protons being being translocated by Red/Ox reactions, this would now cause a build up of protons on the outside of the membrane, separating both charge (positive on the outside and negative on the inside an electrical potential) and pH (low pH outside, higher pH inside). With excess protons on the outside of the cell membrane, and the F0F1ATPase no longer consuming ATP to translocate protons, the pH and charge gradients can be used to drive the F0F1ATPase "backwards" that is to form or produce ATP by using the energy in the charge and pH gradients set up by the Red/Ox pumps as depicted in figure 8 . This arrangement is called an electron transport chain (ETC).

The evolution of the ETC the combination of the Red/Ox driven proton translocators coupled to the production of ATP by the F0F1ATPase.

MODULE 05.7 Pentose Phosphate Pathway

INTRODUCTION In most introductory biology and biochemistry courses focus on glycolysis (oxidation of glucose to pyruvate) and the TCA cycle, the oxidation of pyruvate to acetyl

CoA and the eventual complete oxidation to CO2. While these are extremely important and universal reactions, most courses leave out the pentose phosphate pathway or hexose monophosphate shunt. This pathway, like the TCA cycle is partially cyclic in nature, where 3 glucose molecules enter and 2 glucose and 1 glyceraldyde-3-phosphate leave. The 2 glucose molecules can recycle and the G3P enters glycolysis. Its an important pathway because it is the primary mechanism for the formation of pentoses, the five carbon sugar required for nucleotide biosynthesis as well as the formation of a variety of other essential cellular components and NADPH, the cellular reductant primarily used in anabolic reactions.

A note from the Instuctor As with the modules on glycolysis and the TCA cycle, there is a lot of material in this module. AS with the other modules, I do not expect you to memorize specific names of compounds or enzymes. However, I will give you those names for completeness. For exams I will always provide you with the pathways we discuss in class and in the BioStax Biology text modules. What you need to be able to do is understand what is going on in each reaction. We will go over in lecture, problems that will be similar to those I will ask of you on exams. Do not be overwhelmed with specific enzyme names and specific structures. What you should know are the general types of enzymes used and the types of structures found. For example you do not need to memorize the structures of eyrthose or sedoheptulose. You will need to know that both are sugars, the former a 4-carbon sugar and the latter a 7-carbon sugar. Remember the ending "ose" identifies the compound as a sugar. In addition, you will not need to know the details of the two unique reactins found in the PPP, the transketolase and transaldolase reactions, thow you do need to be able to identify a ketone containing sugar versus an aldehyde containing sugar. Finally, you will not be expected to memorize enzyme names, but like in glycolysis and the TCA cycle you will be expected to know the various types of reactions a type of enzyme can catalyze, for example, a transaldolase moves aldehyde groups from one compound to another. This is the level of understanding I expect. If you have any questions please ask.

Oxidatvie Pentose Phosphate Pathway: AKA The hexose monophosphate shunt

While glycolysis has evolved to oxidize hexoses to form carbon precursors for biosynthesis, energy (ATP) and reducing power (NADH) the Pentose Phosphate Pathway (PPP) has evolved to utilize pentoses or five carbon sugars. Pentose are required precursors for nucleotides and other essential biomolecules. The PPP also generates NADPH instead of NADH, which is required for most anabolic reactions. The PPP, in conjuction with Glycolysis and the TCA cycle make up what we call Central Metabolism. These 3 central pathways (along with the reaction Pyruvate to Acetyl

CoA) are responsible for producing all of the necessary precursor molecules required by all cells. The PPP is responsible for producing pentos-phosphates (5 carbon sugars), Eyrthrose-phosphate (four carbon sugars)and NADPH . This pathway is also responsible for the production of Sedoheptulose -phosphate , an essential 7-carbon sugar used in the outer cell membranes of Gram-negative bacteria.

Below is a diagram of the pathway. The pathway is complex and involves a variety of novel rearrangement reactions that move two and three carbon units around. These reactions called transaldolase and transketalase are used to produce the intermediates within the pathway. The net result is oxidation and subsequent decarboxylation of glucose to form a pentose. The total reaction involves 3 glucose-6-Phosphate (in green) molecules being oxidized to form 3 CO2 molecules, 1 glyceraldehyde-Phosphate (in red), and 2 hexose-phosphates (in red). In this cycle, the formed glyceradehyde-Phosphate feeds into glycolysis and the 2 hexose-Phosphates (glucose-Phosphates) can recycle into the PPP or gycolysis.

Pentose Phosphate Pathway Take home message

As shown in Figure 2, the net result of the pathway is 1 trios-phosphate (glyceraldehyde-3-Phosphate) that can then be further oxidized via glycolysis 2 recycled hexose-phosphates (in the form of either glucose-6-phosphae or fructose-6-phosphate) and NADPH which is required reductant for many biosynthetic (anabolic) reactions. The pathway provides a variety of intermediate sugar-phosphates that the cell may require, such as pentose-phosphates (for nucleotides and some amino acids), erythrose-phosphate (for amino acids) and sedohepulose-phosphate, for Gram-negative bacteria.

The PPP along with glycolysis, the TCA cycle and the oxidation of Pyruvate to acetyl-Co makes up the major pathways of central metabolism and is required to some degree of all organisms to construct the basic substrates to create the building blocks of life.


By the end of this module you should be able to describe the role the pentose phosphate pathway plays in central metabolism. Determine the end-products of the pathway.

MODULE 05.8 Photosynthesis and the Calvin Cycle

INTRODUCTION The light dependent reactions of photosynthesis couple the transfer of energy in light into chemical compounds through a series of redox reactions in an electron transport chain (review module 5.6). In the light dependent reactions, both ATP and NADPH are generated. Using the energy carriers formed in the first steps of photosynthesis, the light-independent reactions, or the Calvin cycle, take in CO2 from the environment and incorporate it into larger biomolecules. An enzyme, RuBisCO, catalyzes a reaction with CO2 and another molecule, RuBP to produce two three carbon sugars. This process is called carbon fixation. After three cycles, a three-carbon molecule of glyceraldehyde-3-phosphate (G3P), the same one we saw earlier in glycolysis, leaves the cycle to become part of a carbohydrate molecule. The remaining G3P molecules stay in the cycle to be regenerated into RuBP, which is then ready to react with more CO2. Photosynthesis forms an energy cycle with the process of cellular respiration. Plants need both photosynthesis and respiration for their ability to function in both the light and dark, and to be able to interconvert essential metabolites. Therefore, plants contain both chloroplasts and mitochondria - more on these organelles soon.

Light Energy

The sun emits an enormous amount of electromagnetic radiation (solar energy). Scientists differentiate the various types of radiant energy from the sun within the electromagnetic spectrum. The electromagnetic spectrum is the range of all possible frequencies of radiation. The human eye can only perceive a small fraction of this energy and this portion of the electromagnetic spectrum is therefore referred to as “visible light.” Visible light constitutes only one of the many types of electromagnetic radiation emitted from the sun and other stars.

The sun emits energy in the form of electromagnetic radiation. This radiation exists at different wavelengths, each of which has its own characteristic energy. All electromagnetic radiation, including visible light, is characterized by its wavelength. Each type of electromagnetic radiation travels at a particular wavelength. The longer the wavelength (or the more stretched out it appears in the diagram), the less energy it carries. Short, tight waves carry the most energy. This may seem illogical, but think of it in terms of a piece of moving a heavy rope. It takes little effort by a person to move a rope in long, wide waves. To make a rope move in short, tight waves, a person would need to apply significantly more energy.

Light Absorption

Light energy initiates many light dependent biological process when pigments absorb photons of light. Organic pigments, whether in the human retina, chloroplast thylakoid, or microbial membrane often have specific ranges of energy levels or wavelengths that they can absorb that are dependent on their chemical makeup and structure. A pigment like the retinal in our eyes, when coupled with an opsin sensor protein, “sees” (absorbs) light predominantly with wavelengths between 700 nm and 400 nm. Because this range defines the physical limits for what light we can see, we refer to it, as noted above, as the "visible range". For similar reasons, plants pigment molecules absorb only light in the wavelength range of 700 nm to 400 nm plant physiologists refer to this range for plants as photosynthetically active radiation.

The visible light seen by humans as white light is composed of a rainbow of colors, each with a characteristic wavelength. Certain objects, such as a prism or a drop of water, disperse white light to reveal the colors to the human eye. In the visible spectrum, violet and blue light have shorter (higher energy) wavelengths while the orange and red light have longer (lower energy) wavelengths.([link]).

The colors of visible light do not carry the same amount of energy. Violet has the shortest wavelength and therefore carries the most energy, whereas red has the longest wavelength and carries the least amount of energy. (credit: modification of work by NASA)

Understanding Pigments

Chlorophylls (including bacteriochlorophylls) and carotenoids are the two major classes of photosynthetic lipid derived pigments found in bacteria, plants and algae each class has multiple types of pigment molecules. There are five major chlorophylls: a, b, c, d, and f. Chlorophyll a is related to a class of more ancient molecules found in bacteria called bacteriochlorophylls . Carotenoids are also very ancient molecules, found in bacteria and eukaryotes. They are the red/orange/yellow pigments found in nature. They are found in fruit—such as the red of tomato (lycopene), the yellow of corn seeds (zeaxanthin), or the orange of an orange peel (β-carotene)—which are used as advertisements to attract seed dispersers (animals or insects that may carry seeds elsewhere). In photosynthesis, carotenoids function as photosynthetic pigments. In addition, when a leaf is exposed to full sun, that surface is required to process an enormous amount of energy if that energy is not handled properly, it can do significant damage. Therefore, many carotenoids help absorb excess energy in light and safely dissipate that energy as heat.

Each type of pigment can be identified by the specific pattern of wavelengths it absorbs from visible light, which is the absorption spectrum . The graph in [link] shows the absorption spectra for chlorophyll a, chlorophyll b, and a type of carotenoid pigment called β-carotene (which absorbs blue and green light). Notice how each pigment has a distinct set of peaks and troughs, revealing a highly specific pattern of absorption. Chlorophyll a absorbs wavelengths from either end of the visible spectrum (blue and red), but not green. Because green is reflected or transmitted, chlorophyll appears green. Carotenoids absorb in the short-wavelength blue region, and reflect the longer yellow, red, and orange wavelengths.

(a) Chlorophyll a, (b) chlorophyll b, and (c) β-carotene are hydrophobic organic pigments found in the thylakoid membrane. Chlorophyll a and b, which are identical except for the part indicated in the red box, are responsible for the green color of leaves. β-carotene is responsible for the orange color in carrots. Each pigment has (d) a unique absorbance spectrum.

Many photosynthetic organisms have a mixture of pigments which optimizes the organism's ability to absorb energy from a wider range of wavelengths. Not all photosynthetic organisms have full access to sunlight. Some organisms grow underwater where light intensity and available wavelengths decrease and change, respectively, with depth. Other organisms grow in competition for light. Plants on the rainforest floor must be able to absorb any bit of light that comes through, because the taller trees absorb most of the sunlight and scatter the remaining solar radiation ([link]).

Plants that commonly grow in the shade have adapted to low levels of light by changing the relative concentrations of their chlorophyll pigments. (credit: Jason Hollinger)

Photophosphorylation an Overview:

Photophosphorylation is the process of transferring the energy from light into chemicals, in particular ATP and NADPH. The evolutionary roots of photophosphorylation are likely in the anaerobic world, between 3 billion and 1.5 billion years ago, when life was abundant in the absence of molecular oxygen. Photophosphorylation probably evolved relatively shortly after electron transport chains and anaerobic respiration began to provide metabolic diversity. The first step of the process involves the absorption of a photon by a pigment molecule. Light energy is transferred to the pigment and promotes electrons into a higher potential energy state - termed an "excited state". The electrons are colloquially said to be "energized". In the excited state, the pigment now has a very low reduction potential and can donate the "excited" electrons to other carriers with greater reduction potentials. These electron acceptors may in turn become donors to other molecules with greater reduction potentials and in so doing form an electron transport chain. As electrons pass from one electron carrier to another via red/ox reactions, these exergonic transfers can be coupled to the endergonic transport (or pumping) of protons across a membrane to create an electrochemical gradient. This electrochemical gradient generates a proton motive force whose exergonic drive to reach equilibrium can be coupled to the endergonic production of ATP, via ATP synthase. As we will seen in more detail, the electrons involved in this electron transport chain can have one of two fates: (1) they may be returned to their initial source in a process called cyclic photophosphorylation or (2) they can be deposited onto a close relative of NAD + called NADP + . If the electrons are deposited back on the original pigment in a cyclic process, the whole process can start over. If, however, the electron is deposited onto NADP + to form NADPH (**shortcut note - we didn't explicitly mention any protons but assume it is understood that they are also involved**) the original pigment must regain an electron from somewhere else. This electron must come from a source with a smaller reduction potential than the oxidized pigment and depending on the system there are different possible sources, including H2O, reduced sulfur compounds such as SH2 and even elemental S 0 .

What happens when a compound absorbs a photon of light?

When a compound absorbs a photon of light, the compound is said to leave its ground state and become "excited", in the sense that it has this extra energy. This is illustrated in figure 5 schematically.

A diagram of what happens to a molecule that absorbs a photon of light.

What are the fates of the "excited" electron? There are four possible outcomes, which are schematically diagrammed in Figure 6 below. These options are:

  1. The electron can relax to a lower orbital, transferring energy as heat.
  2. The electron can relax to a lower orbital and transfer energy into a photon of light - a process known as fluorescence .
  3. The energy can be transferred by resonance to a neighboring molecule as the e - returns to a lower orbital.
  4. The energy can change the reduction potential such that the molecule can become an e - donor. Linking this excited e - donor to a proper e - acceptor can lead to an exergonic electron transfer. In other words, the excited state can be involved in Red/Ox reactions.

As the excited electron decays back to its original orbit, the energy can be transferred in a variety of ways. While many so called antenna or auxiliary pigments absorb light energy and transfer it to something known as a reaction center (by mechanisms depicted in option III in figure 6) it is what happens at the reaction center that we are most concerned with (option IV in figure 6). Here a chlorophyll or bacteriochlorophyll molecule absorbs a photon's energy and an electron is excited. This energy transfer is sufficient to allow the reaction center to donate the electron in a red/ox reaction to a second molecule. This initiates the photophosphorylation electron transport reactions. The result is an oxidized reaction center that must now be reduced in order to start the process again. How this happens is the basis of electron flow in photophosphorylation and will be described in detail below.

Simple Photophosphorylation Systems: Anoxygenic photophosphorylation

Introduction Early in the evolution of photophosphorylation, these reactions evolved in anaerobic environments where there was very little molecular oxygen available. Two sets of reactions evolved under these conditions, both directly from anaerobic respiratory chains as described above. These are known as the light reactions because they require the activation of an electron (an excited electron) from the absorption of a photon of light by a reaction center pigment, such as bacteriochlorophyll. The light reactions are categorized either as cyclic or as noncyclic photophosphorylation, depending upon the final state of the electron(s) removed from the reaction center pigments. If the electron(s) return to the original pigment reaction center, such as bacteriochlorophyll, this is cyclic photophosphorylation the electrons make a complete circuit and is diagramed in figure 8. If the electron(s) are used to reduce NADP + to NADPH, the electron(s) are removed from the pathway and end up on NADPH, this process is referred to as noncyclic since the electrons are no longer part of the circuit. In this case the reaction center must be re-reduced before the process can happen again. Therefore, an external electron source is required for noncylic photophosphorylation. In these systems reduced forms of Sulfur, such as H2S, which can be used as an electron donor and is diagrammed in figure 9. To help you better understand the similarities of photophosphorylation to respiration, a redox tower (figure 7) has been provided that contains many commonly used compounds involved with photosphosphorylation.

Electron tower that has a variety of common photophosphorylation components. PSI and PSII refer to Photosystems I and II of the oxygenic photophosphorylation pathways. For the examples in Figure 8 and Figure 9 P840 is similar in reduction potential as is PSI.

Cyclic Photophosphorylation In cyclic photophosphorylation the bacteriochlorophyllred molecule absorbs enough light energy to energize and eject an electron to form bacteriochlorophyllox. The electron reduces a carrier molecule in the reaction center which in turn reduces a series of carriers via red/ox reactions. These carriers are the same carriers found in respiration. If the change in reduction potential from the various red/ox reactions are sufficiently large, protons, H + are translocated across the membrane. Eventually the electron is used to reduce bacteriochlorophyllox and the whole process can start again. This is called cyclic photophosphorylation because the electrons make a complete circuit: bacteriochlorophyll is the source of electrons and is the final electron acceptor. ATP is produced via the F1F0 ATPase . The schematic in figure 8 below demonstrates how cyclic photophosphorylation works.

Cyclic Photophosphorylation. The reaction center P840 absorbs light energy and becomes excited, denoted with an *. The excited electron is ejected and used to reduce an FeS protein leaving an oxidized reaction center. The electron its transferred to a quinone, then to a series of cytochromes which in term is reduces the P840 reaction center. The process is cyclical. Note the gray array coming from the FeS protein going to a ferridoxin (Fd), also in gray. This represents an alternative pathway the electron can take and will be discussed below in non-cyclic photophosphorylation. NOTE the same electron that leaves the P480 reaction center is not necessarily the same electron that eventually finds its way back to reduce the oxidized P840.

Non-cyclic photophosphorylation In cyclic photophosphorylation electrons cycle from bacteriochlorophyll (or chlorophyll) to a series of electron carriers and eventually back to bacteriochlorophyll (or chlorophyll): there is no loss of electrons, they stay in the system. In non-cyclic photophosphorylation the electrons are removed from the photosystem and redox chain and they eventually end up on NADPH. That means there needs to be a source of electrons, a source that has a smaller reduction potential than bacteriochlorophyll (or chlorophyll) that can donate electrons to bacteriochlorophyllox to reduce it. An electron tower is proved above so you can see what compounds can be used to reduce the oxidized form of bacteriochlorophyll. The second requirement, is that when bacteriochlorophyll becomes oxidized and the electron is ejected it must reduce a carrier that has a greater reduction potential than NADP/NADPH (see the electron tower). In this case, electrons can flow from energized bacteriochlorophyll to NADP forming NADPH and oxidized bacteriochlorophyll. Electrons are lost from the system and end up on NADPH, to complete the circuit bacteriochlorophyllox is reduced by an external electron donor, such as H2S or elemental S 0 .

Non-cyclic photophosphorylation. In this example, the P840 reaction center absorbs light energy and becomes energized, the emitted electron reduced a FeS protein and in turn reduces ferridoxin. Reduced ferridoxin (Fdred) can now reduce NADP to form NADPH. The electrons are now removed from the system, finding their way to NADPH. The electrons need to be replaced on P840, which requires an external electron donor. In this case, H2S serves as the electron donor.

Possible Discussion It should be noted that for bacterial photophosphorylation pathways, for each electron donated from a reaction center (remember only one electron is actually donated/reaction center (or chlorophyl molecule), the resulting output from that electron transport chain is either the formation of NADPH (requires 2 electrons)or ATP can be made, NOT not both. In other words, the path the electrons take in the ETC can have one or two possible outcomes. This puts limits on the versatility of the bacterial anoxygenic photosynthetic systems. But what would happen if there evolved a process that utilized both systems, that is a cyclic and non-cyclic photosynthetic pathway? That is, if both ATP and NADPH could be formed from from a single input of electrons? A second limitation is that these bacterial systems require compounds such as reduced sulfur to act as electron donors to reduce the oxidized reaction centers, not necessarily widely found compounds. What would happen if a chlorophyllox molecule would have a reduction potential higher (more positive) than that of the molecular the O2/H2O reaction? Answer, a planetary game changer.

Oxygenic Photophosphorylation

Generation of NADPH and ATP The overall function of light-dependent reactions is to transfer solar energy into chemical compounds, largely the the molecules NADPH and ATP. This energy supports the light-independent reactions and fuels the assembly of sugar molecules. The light-dependent reactions are depicted in [link]. Protein complexes and pigment molecules work together to produce NADPH and ATP.

A photosystem consists of a light-harvesting complex and a reaction center. Pigments in the light-harvesting complex pass light energy to two special chlorophyll a molecules in the reaction center. The light excites an electron from the chlorophyll a pair, which passes to the primary electron acceptor. The excited electron must then be replaced. In (a) photosystem II, the electron comes from the splitting of water, which releases oxygen as a waste product. In (b) photosystem I, the electron comes from the chloroplast electron transport chain discussed below.

The actual step that transfers light energy into the biomolecule takes place in a multiprotein complex called a photosystem , two types of which are found embedded in the thylakoid membrane, photosystem II (PSII) and photosystem I (PSI) ([link]). The two complexes differ on the basis of what they oxidize (that is, the source of the low-energy electron supply) and what they reduce (the place to which they deliver their energized electrons).

Both photosystems have the same basic structure a number of antenna proteins to which the chlorophyll molecules are bound surround the reaction center where the photochemistry takes place. Each photosystem is serviced by the light-harvesting complex , which passes energy from sunlight to the reaction center it consists of multiple antenna proteins that contain a mixture of 300� chlorophyll a and b molecules as well as other pigments like carotenoids. The absorption of a single photon or distinct quantity or “packet” of light by any of the chlorophylls pushes that molecule into an excited state. In short, the light energy has now been captured by biological molecules but is not stored in any useful form yet. The energy is transferred from chlorophyll to chlorophyll until eventually (after about a millionth of a second), it is delivered to the reaction center. Up to this point, only energy has been transferred between molecules, not electrons.

In the photosystem II (PSII) reaction center, energy from sunlight is used to extract electrons from water. The electrons travel through the chloroplast electron transport chain to photosystem I (PSI), which reduces NADP + to NADPH. The electron transport chain moves protons across the thylakoid membrane into the lumen. At the same time, splitting of water adds protons to the lumen, and reduction of NADPH removes protons from the stroma. The net result is a low pH in the thylakoid lumen, and a high pH in the stroma. ATP synthase uses this electrochemical gradient to make ATP.

The reaction center contains a pair of chlorophyll a molecules with a special property. Those two chlorophylls can undergo oxidation upon excitation they can actually give up an electron in a process called a photo activation . It is at this step in the reaction center, this step in photosynthesis, that light energy is transferred into an excited electron. All of the subsequent steps involve getting that electron onto the energy carrier NADPH for delivery to the Calvin cycle where the electron is deposited onto carbon for long-term storage in the form of a carbohydrate. PSII and PSI are two major components of the photosynthetic electron transport chain , which also includes the cytochrome complex . The cytochrome complex, an enzyme composed of two protein complexes, transfers the electrons from the carrier molecule plastoquinone (Pq) to the protein plastocyanin (Pc), thus enabling both the transfer of protons across the thylakoid membrane and the transfer of electrons from PSII to PSI.

The reaction center of PSII (called P680 ) delivers its high-energy electrons, one at a time, to the primary electron acceptor , and through the electron transport chain (Pq to cytochrome complex to plastocyanine) to PSI. P680’s missing electron is replaced by extracting an electron from water thus, water is split and PSII is re-reduced after every photoact. Splitting one H2O molecule releases two electrons, two hydrogen atoms, and one atom of oxygen. Splitting two molecules of water is required to form one molecule of diatomic O2 gas. About 10 percent of the oxygen is used by mitochondria in the leaf to support oxidative phosphorylation. The remainder escapes to the atmosphere where it is used by aerobic organisms to support respiration.

As electrons move through the proteins that reside between PSII and PSI, they take part in exergonic redox transfers. The free energy associated with the exergonic redox reaction is coupled to the endergonic transport of hydrogen atoms from the stromal side of the membrane to the thylakoid lumen. Those hydrogen atoms, plus the ones produced by splitting water, accumulate in the thylakoid lumen and will be used to synthesize ATP in a later step. Since the electrons on PSI now have a greater reduction potential than when they started their trek (it is important to note that PSI unexcited sits lower on the redox tower than NADP+/NADPH), they must be re-energized in PSI. Therefore, another photon is absorbed by the PSI antenna. That energy is transferred to the PSI reaction center (called P700 ). P700 is oxidized and sends an electron through several intermediate redox steps to NADP + to form NADPH. Thus, PSII captures the energy in light and couples its transfer via redox reactions to the creation of a proton gradient. The exergonic and controlled relaxation of this gradient can be coupled to the synthesis of ATP. PSI captures energy in light and couples that, through a series of redox reactions, to reduce NADP + into NADPH. The two photosystems work in concert, in part, to guarantee that the production of NADPH will be in the right proportion to the production of ATP. Other mechanisms exist to fine tune that ratio to exactly match the chloroplast’s constantly changing energy needs.

Additional links

Light Independent Reactions or Carbon Fixation

A short introduction The general principle of carbon fixation is that some cells under certain conditions can take inorganic carbon, CO2 (also referred to as mineralized carbon) and reduce it to a usable cellular form. Most of us are aware that green plants can take up CO2 and produce O2 in a process known as photosynthesis. We have already discussed photophosphorylation, the ability of a cell to transfer light energy onto chemicals and ultimately to produce the energy carriers ATP and NADPH in a process known as the light reactions. In photosynthesis, the plant cells use the ATP and NADPH formed during photophosphorylation to reduce CO2 to sugar, (as we will see, specifically G3P) in what are called the dark reactions. While we appreciate that this process happens in green plants, photosynthesis had its evolutionary origins in the bacterial world. In this module we will go over the general reactions of the Calvin Cycle, a reductive pathway that incorporates CO2 into cellular material.

In photosynthetic bacteria, such as Cyanobacteria and purple non-sulfur bacteria, as well plants, the energy (ATP) and reducing power (NADPH) - a term used to describe electron carriers in their reduced state - obtained from photophosphorylation is coupled to " Carbon Fixation ", the incorporation of inorganic carbon (CO2) into organic molecules initially as glyceraldehyde-3-phosphate (G3P) and eventually into glucose. Organisms that can obtain all of their required carbon from an inorganic source (CO2)are refereed to as autotrophs , while those organisms that require organic forms of carbon, such as glucose or amino acids, are refereed to as heterotrophs . The biological pathway that leads to carbon fixation is called the Calvin Cycle and is a reductive pathway (consumes energy/uses electrons) which leads to the reduction of CO2 to G3P.

The Calvin Cycle: the reduction of CO2 to Glyceraldehyde 3-Phosphate

Light reactions harness energy from the sun to produce chemical bonds, ATP, and NADPH. These energy-carrying molecules are made in the stroma where carbon fixation takes place.

In plant cells, the Calvin cycle is located in the chloroplasts. While the process is similar in bacteria, there are no specific organelles that house the Calvin Cycle and the reactions occur in the cytoplasm around a complex membrane system derived from the plasma membrane. This intracellular membrane system can be quite complex and highly regulated. There is strong evidence that supports the hypothesis that the origin of chloroplasts from a symbiosis between cyanobacteria and early plant cells.

Stage 1: Carbon Fixation

In the stroma of plant chloroplasts, in addition to CO2, two other components are present to initiate the light-independent reactions: an enzyme called ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), and three molecules of ribulose bisphosphate (RuBP), as shown in [link]. Ribulose-1,5-bisphosphate (RuBP) is composed of five carbon atoms and includes two phosphates.

The Calvin cycle has three stages. In stage 1, the enzyme RuBisCO incorporates carbon dioxide into an organic molecule, 3-PGA. In stage 2, the organic molecule is reduced using electrons supplied by NADPH. In stage 3, RuBP, the molecule that starts the cycle, is regenerated so that the cycle can continue. Only one carbon dioxide molecule is incorporated at a time, so the cycle must be completed three times to produce a single three-carbon GA3P molecule, and six times to produce a six-carbon glucose molecule.

RuBisCO catalyzes a reaction between CO2 and RuBP. For each CO2 molecule that reacts with one RuBP, two molecules of another compound (3-PGA) form. PGA has three carbons and one phosphate. Each turn of the cycle involves only one RuBP and one carbon dioxide and forms two molecules of 3-PGA. The number of carbon atoms remains the same, as the atoms move to form new bonds during the reactions (3 atoms from 3CO2 + 15 atoms from 3RuBP = 18 atoms in 3 atoms of 3-PGA). This process is called carbon fixation , because CO2 is “fixed” from an inorganic form into an organic molecule.

Stage 2: Reduction

ATP and NADPH are used to convert the six molecules of 3-PGA into six molecules of a chemical called glyceraldehyde 3-phosphate (G3P) - a carbon compound that also is produced in glycolysis. Six molecules of both ATP and NADPH are used in the process. The exergonic process of ATP hydrolysis is in effect driving the endergonic redox reactions, creating ADP and NADP + . Both of these molecules return to the nearby light-dependent reactions to be recycled back into ATP and NADPH.

Stage 3: Regeneration

Interestingly, at this point, only one of the G3P molecules leaves the Calvin cycle to contribute to the formation of other compounds needed by the organism. In plants, because the G3P exported from the Calvin cycle has three carbon atoms, it takes three “turns” of the Calvin cycle to fix enough net carbon to export one G3P. But each turn makes two G3Ps, thus three turns make six G3Ps. One is exported while the remaining five G3P molecules remain in the cycle and are used to regenerate RuBP, which enables the system to prepare for more CO2 to be fixed. Three more molecules of ATP are used in these regeneration reactions.

The harsh conditions of the desert have led plants like these cacti to evolve variations of the light-independent reactions of photosynthesis. These variations increase the efficiency of water usage, helping to conserve water and energy. (credit: Piotr Wojtkowski)

Free Response Questions

Why is the third stage of the Calvin cycle called the regeneration stage?

Because RuBP, the molecule needed at the start of the cycle, is regenerated from G3P.

Which part of the light-independent reactions would be affected if a cell could not produce the enzyme RuBisCO?

None of the cycle could take place, because RuBisCO is essential in fixing carbon dioxide. Specifically, RuBisCO catalyzes the reaction between carbon dioxide and RuBP at the start of the cycle.

Why does it take three turns of the Calvin cycle to produce G3P, the initial product of photosynthesis?

Because G3P has three carbon atoms, and each turn of the cycle takes in one carbon atom in the form of carbon dioxide.

Prepare for the Test: Create an energy story for each phase of the Calvin cycle. Classify the reactants and products and pay attention to where the energy is at the beginning of the reaction and the end of the reaction. At this point you should be able to tell if a reaction is a REDOX reaction (does it have NADPH as a reactant or product?) or if the reaction is endergonic or exergonic (is ATP created or used in the reaction?).

Synthesis of Nucleotides

Purines (Adenine & Guanine) and pyrimidines (Thymine, Cytosine & Uracil) are the two classes of nucleotides which forms the nucleic acids (DNA & RNA) in the cells. Apart from the primary role of DNA and RNA as “genetic information storage”, nucleotides also serves different functions in the cells such as energy carrier (ATP and GTP), components of co-enzymes (NAD and FAD) and cellular signal transduction (cAMP and cGMP as ‘second messengers’). An ample supply of nucleotides in the cell is very essential for all the cellular processes. This post discuss the biosynthesis of Purines and Pyrimidines in an EASY but detailed way.

Pathways for the biosynthesis of nucleotides

Nucleotide biosynthesis in the cell can be grouped into two broad classes. (1) de-novo synthesis and (2) synthesis by salvage pathways.

I. De-novo synthesis (synthesis from scratch): it is a biochemical pathway in which nucleotides are synthesized new from simple precursor molecules.
II. Salvage pathway (recycle pathway): used to recover bases and nucleosides formed during the degradation of RNA and DNA

Learning objectives:

@. How nucleotides are synthesized in the cells?
@. How de-novo synthesis of purines & pyrimidines occurs?
@. Synthesis of IMP (precursor of Adenine and Guanine)
@. Synthesis of Adenine and Guanine from IMP
@. Synthesis of Uracil
@. Synthesis of Cytosine
@. Synthesis of deoxyribonucleotides
@. Synthesis of Thymine
@. Salvage pathways of purines and pyrimidines

. I. De-novo synthesis of purines:

The purine nucleotides of nucleic acids are adenosine 5-monophosphate (AMP adenylate) and guanosine 5-monophosphate (GMP guanylate), containing the purine bases adenine and guanine respectively. The first idea about purine nucleotide biosynthesis in the cell was come from the study of John Buchanan (1948) by radioactive tracer studies in birds by analyzing the biochemistry of uric acid (a purine present in the excreta of birds). The detailed biosynthetic pathways of the purine biosynthesis came latter in 1950 primarily by the works of Buchanan and G. Robert Greenberg.

The image shows the source of different atoms in a purine skeleton identified by radio labeling studies

N1 is derived from amino group of Aspartate

C2 & C8 is derived from Formate

N3 & N9 is derived from amide group of Glutamine

C4, C5 & N7 is derived from Glycine

C6 is derived from HCO3- (bicarbonate)

Aspartate, Formate, Glutamine, Glycine and Bicarbonate acts as the building blocks for purine synthesis

Purines (adenine and guanine) are synthesized as ribo-nucleotides (nitrogen base + ribose sugar + phosphate) rather than as free bases. Both purines are derived from a precursor namely inosine-5′-monophosphate (IMP). Thus the purine synthesis starts with IMP synthesis (See the mind map) .

Synthesis of Inosine monophosphate (IMP):

Inosine monophosphate (IMP) is synthesized in 11 enzymatic steps from simple precursors as summarized below

Step-1: Ribose-5-phosphate activation and formation of PRPP): α-D-Ribose-phosphate (R5P) is activated with ATP to form 5-phosphoribosyl-α-pyrophosphate (PRPP) with the help of enzyme Ribose phosphate pyrophosphokinase. PRPP is also one of the precursors for the synthesis of pyrimidines and also the amino acids Histidine and Tryptophan.

Step-2: Acquisition of N9 atom of purine: Amide nitrogen of glutamine displaces the pyrophosphate group of PRPP and it also inverts the configuration at C1′ to form β-5-phosphoribosylamine (PRA) with the help of enzyme amidophosphoribozyl transferase. (This reaction contribute N9 atom of purine form glutamine)

Step-3: Acquisition of C4, C5 & N7 atoms of purine: Carboxylic group of glycine is combined with the amino group of β-5-phosphoribosylamine (PRA) to form glycinamide ribotide (GAR) with the help of enzyme – GAR synthetase (C4, C5, & N7 of purine are contributed by glycine)

Step-4: Acquisition of C8 atom of purine: Amino group of glycinamide ribotide (GAR) is formylated with N10-formyltetrahydrofolate and forms formylglycinamide ribotide (FGAR) with the presence of enzyme GAR transformylase. (C8 of purine is contributed by formate)

Step-5: Acquisition of N3 atom of purine: Amide nitrogen of second glutamine is added to FGAR in an ATP-dependent reaction to form formylglycinamidine ribotide (FGAM) with the help of enzyme FGAM synthetase. (N3 of purine is contributed by glutamine)

Step-6: Purine imidazole ring formation: An ATP dependent ring closing (imidazole ring formation) reaction in the presence of AIR synthetase enzyme to produce 5-aminoimidazole ribotide (AIR).

Step-7: Acquisition of C6 atom of purine: An ATP dependent carboxylation reaction of 5-aminoimidazole ribotide (AIR) with HCO3- (bicarbonate) to produce carboxyaminoimidazole ribotide (CAIR) in the presence of enzyme AIR carboxylase. (C6 of purine is contributed by HCO3-)

Step-8: Acquisition of N1 atom of purine: Aspartate is added and it forms an amide bond with C6 to form 5-aminoimidazole-4-(N-succinylocarboxamide) ribotide (SACAIR) in an ATP dependent reaction with the help of enzyme SAICAR synthetase (N1 of purine is contributed by aspartate)

Step-9: Elimination of fumarate: Fumarate group is cleaved off from SACAIR to produce 5-aminoimidazole-4-carboxamide ribotide (AICAR) with the help of enzyme- adenylosuccinate lyase.

Step-10: Acquisition of C2 atom of purine: Amino group of AICAR react with N10-formyltetrahydrofolate (formylation) to form 5-formaminoimidazole-4-carboxamide ribotide (FAICAR) with presence of enzyme AICAR transformylase. (C2 of purine ring is contributed by this N10-formyltetrahydrofolate)

Step-11: Cyclization to form IMP: In the last reaction, the larger ring of FAICAR is enzymatically closed to forms Inosine Monophosphate (IMP) with the release of a water molecule catalyzed by the enzyme IMP cyclohydrolase

Synthesis of Adenine and Guanine

IMP does not accumulate in the cells rather it is rapidly converted into Adenine (as AMP) and Guanine (as GMP). AMP differ from IMP in the replacement of its 6-keto group by an amino group whereas GMP differ from IMP in the presence of an amino group at C2

(a). Synthesis of AMP (Adenosine Monophosphate)

IMP is converted to AMP in two enzymatic steps

Step-1: Donation of amino group by aspartate: Amino group of aspartate is enzymatically linked to the IMP (C6 of purine) coupled with GTP hydrolysis to form adenylosuccinate with the help of enzyme- adenylosuccinate synthetase.

Step-2: Eliminates fumarate group to form AMP: Adenylosuccinate is enzymatically converted to AMP by the removal of fumarate group with the help of enzyme adenylosuccinate lyase.

(b). Synthesis of GMP (Guanosine Monophosphate)

IMP is converted to GMP in two enzymatic steps

Step-1: Dehydrogenation of IMP: IMP is enzymatically dehydrogenated to form Xanthosine Monophosphate (XMP) with the enzyme IMP dehydrogenase. The H+ ions released are accepted by NAD+.

Step-2: Amidation of XMP: In the second step, XMP is amidated with the amide group from glutamine with the presence of H2O and hydrolysis of ATP yields GMP (Guanosine monophosphate) catalyzed by the enzyme GMP synthetase.

Synthesis of Nucleoside Diphosphates and Triphosphates

For the participation of DNA and RNA synthesis, nucleoside monophosphates and diphosphates must be converted into nucleoside triphosphates. Nucleotide diphosphates are synthesized from corresponding nucleotide monophosphate by phosphate group transfer from ATP with the help of base specific nucleoside monophosphate kinase enzyme. Similarly, nucleotide triphosphates are synthesized by the second round phosphorylation aided by ATP with the help of enzyme nucleoside diphosphate kinase.

ADP can also be converted to ATP by various energy-releasing reactions in the cells such as by oxidative phosphorylation (electron transport system of respiration), by photophosphorylation (light reaction of photosynthesis) and also by substrate level phosphorylation (as in glycolysis)

II. De-novo synthesis of Pyrimidines (Uracil, Thymine & Cytosine)

Biosynthesis of pyrimidines is simple than that of purines. Unlike purine synthesis, pyrimidines are synthesized as bases and latter it is added to ribose sugar, i.e., the ring is completed before being it is linked to ribose-5-phosphate. Following diagram shows the source of different atoms in a pyrimidine skeleton identified by radio labeling studies.

N1, C6, C5 and C4 are derived from aspartate

N3 is derived from glutamine

C2 is derived from HCO3- (bicarbonate)

Aspartate, Glutamine and bicarbonate contributes pyrimidine nucleus

(a). De-novo synthesis of UMP (Uridine monophosphate)

Uridine monophosphate (UMP) also acts as the precursor of CTP and dTTP). De-novo synthesis of UMP is completed in 6 enzymatic steps from simple precursors.

Step-1: Synthesis of carbamoyl phosphate: With the hydrolysis of two ATP molecules, bicarbonate and amide nitrogen of glutamine combine to form carbamoyl phosphate in the presence of enzyme carbamoylphosphate synthetase II.

Step-2: Synthesis of carbamoyl aspartate: Carbamoyl phosphate reacts with aspartate to yield carbamoyl aspartate catalyzed by the enzyme aspartate transcarbamoylase (ATCase).

Step-3: Ring closure & dihydroorotate formation: By the elimination (condensation reaction) of one molecule of water, the carbamoyl aspartate is converted to a ring compound – dihydroorotate catalyzed by dihydroorotase enzyme.

Step-4: Oxidation of dihydroorotate: Dihydroorotate is dehydrogenated to form orotate with the enzyme dihydroorotate dehydrogenase. (In eukaryotes, dihydroorotate dehydrogenase is located in the outer surface of inner mitochondrial membrane. All other enzymes of pyrimidine synthesis are located in the cytosol. Inhibition of dihydroorotate dehydrogenase will inhibit pyrimidine synthesis in T lymphocytes, thereby it attenuate the autoimmune disease rheumatoid arthritis. Since the enzyme is not in the cytosol, the oxidizing power required for the conversion of dihydroorate is provided by Quinone)

Step-5: Acquisition of the ribose phosphate moiety: Orotate reacts with PRPP to produce orotidine-5′-monophosphate (OMP) with the enzyme orotate phosphoribosyl transferase. The anomeric form of pyrimidine nucleotides is fixed in in the β-configuration.

Step-6: Decarboxylation to form UMP: OMP undergoes decarboxylation with assistance of enzyme OMP decarboxylase (ODCase) to form uridine monophosphate (UMP). The catalytic conversion rate of OMP decarboxylase is by a factor of 2 X 1023 over un-catalyzed reaction, making it the most catalytically proficient enzyme known to science.

Synthesis of UTP from UMP

UMP is converted to UTP in two step kinase reaction with 2 molecules of ATP

(b). Synthesis of CTP

CTP is synthesized by the amination of UTP by the enzyme CTP synthase. In animals amino group is donated by glutamine whereas in bacteria, the amino group is donated directly by ammonia.

Synthesis of Deoxyribonucleotides:

Deoxyribonucleotides are synthesized from their corresponding ribonucleotides by the reduction of ribose sugar at position C2’. Enzymes the in the formation of deoxyribonucleotides by the reduction of the corresponding ribonucleotides are called ribonucleotide reductases (RNRs). There are three classes of RNRs so far described in the living world and they all differs in their prosthetic groups. All of them replace the C2’ – OH group of ribose with – H via a free radical mechanism

(c). Synthesis of Thymine (5-methyluracil) as dTTP:

Thymine, which is present in DNA and not in RNA, is a methylated uracil residue. Thymine in the cell is synthesized as dTTP from dUMP by methylation in four steps.

Step-1: dUTP is hydrolyzed to dUMP and PPi by the enzyme dUTP diphosphohydrolase (dUTPase)

Step-2: dUMP is then methylated to form dTMP

Step-3 & 4: dTMP is then phosphorylated with ATP in two rounds to form dTTP

Salvage pathway of Purines

Purines can be generated in the cells during the degradation of nucleic acids through salvage pathways. Turnover of nucleic acids (particularly RNA) in most cells releases adenine, guanine, and hypoxanthine. These free purines are reconverted to their corresponding nucleotides through salvage pathways. The salvage pathways are diverse in different organism in contrast to the de-novo purine nucleotide synthetic pathway which is virtually identical in all cells.

Purines are salvaged by two different enzymes in mammals:

1. Adenine phosphoribosyltransferase (APRT) which mediates AMP formation using PRPP

2. Hypoxanthine–guanine phosphoribosyltransferase (HGPRT), which catalyzes the analogous reaction for both hypoxanthine and guanine

Hypoxanthine + PRPP ⇌ IMP + PPi

Lesch–Nyhan Syndrome (an X-linked trait and thus more common in males) is caused by the deficiency of HGPRT. This syndrome results in excessive uric acid (a purine degradation product) production which leads to neurological abnormalities, mental retardation and aggressive and destructive behavior.

Salvage pathway of pyramidines

Similar to purines, pyramidines are also recovered from the derivative intermediates of nucleic acids such as DNA and RNA. The recoveries of pyrimidines are catalyzed by the enzyme pyrimidine phosphoribosyltransferase which utilizes PRPP as the source of ribose-5-phsophate.

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30 Diagram And Explain Electron Transport

Complex i iv each play a role in transporting electrons hence the name electron transport chain and establishing the proton gradient. This is important because the oxidized forms of these electron carriers are used in glycolysis and the citric acid cycle and must be available to keep these processes running.

Oxidative Phosphorylation Biology Article Khan Academy

In the process protons are pumped from the mitochondrial matrix to the intermembrane space and oxygen is reduced to form water.

Diagram and explain electron transport. This electron transport chain only occurs when oxygen is available. The electron transport chain is a cluster of proteins that transfer electrons through a membrane within mitochondria to form a gradient of protons that drives the creation of adenosine triphosphate atp. Molecular oxygen o 2 acts as an electron acceptor in complex iv and gets converted to a water molecule h 2 o.

Explanation of electron transport system in the largest biology dictionary online. Free learning resources for students covering all major areas of biology. Each enzyme complex carries out the transport of electrons accompanied by the release of protons in the intermembrane space.

Electron transport chain definition. The electron transport chain is a series of electron transporters embedded in the inner mitochondrial membrane that shuttles electrons from nadh and fadh 2 to molecular oxygen. This is shown by the diagram below.

Nadh and fadh2 pass their electrons to the electron transport chain turning back into nad and fad. The exact mechanism of each complex can be overwhelming so i will save that for a future post. The electron is then transported to complex ii which brings about the conversion of succinate to fumarate.

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1 Diagram Of Photosystem Ii Ps Ii Representing The Polypeptide

In vivo photosynthetic electron transport does not limit photosynthetic capacity in phosphate-deficient sunflower and maize leaves

The effects of extreme phosphate (Pi) deficiency during growth on the contents of adenylates and pyridine nucleotides and the in vivo photochemical activity of photosystem II (PSII) were determined in leaves of Helianthus annuus and Zea mays grown under controlled environmental conditions. Phosphate deficiency decreased the amounts of ATP and ADP per unit leaf area and the adenylate energy charge of leaves. The amounts of oxidized pyridine nucleotides per unit leaf area decreased with Pi deficiency, but not those of reduced pyridine nucleotides. This resulted in an increase in the ratio of reduced to oxidized pyridine nucleotides in Pi-deficient leaves. Analysis of chlorophyll a fluorescence at room temperature showed that Pi deficiency decreased the efficiency of excitation capture by open PSII reaction centres (φe), the in vivo quantum yield of PSII photochemistry (φPSII) and the photochemical quenching co-efficient (qP), and increased the non-photochemical quenching co-efficient (qN) indicating possible photoinhibitory damage to PSII. Supplying Pi to Pi-deficient sunflower leaves reversed the long-term effects of Pi-deficiency on PSII photochemistry. Feeding Pi-sufficient sunflower leaves with mannose or FCCP rapidly produced effects on chlorophyll a fluorescence similar to long-term Pi-deficiency. Our results suggest a direct role of Pi and photophosphorylation on PSII photochemistry in both long-and short-term responses of photosynthetic machinery to Pi deficiency. The relationship between φPSII and the apparent quantum yield of CO2 assimilation determined at varying light intensity and 21 kPa O2 and 35 Pa CO2 partial pressures in the ambient air was linear in Pi-sufficient and Pi-deficient leaves of sunflower and maize. Calculations show that there was relatively more PSII activity per mole of CO2 assimilated by the Pi-deficient leaves. This indicates that in these leaves a greater proportion of photosynthetic electrons transported across PSII was used for processes other than CO2 reduction. Therefore, we conclude that in vivo photosynthetic electron transport through PSII did not limit photosynthesis in Pi-deficient leaves of sunflower and maize and that the decreased CO2 assimilation was a consequence of a smaller ATP content and lower energy charge which restricted production of ribulose, 1-5, bisphosphate, the acceptor for CO2.

16 December 2013: I have finalized the draft of the final exam. The format is similar to previous final exams. You will be asked to answer FIVE of the SIX questions in the exam booklet provided.

27 November 2013: I've mounted the lecture notes for the Dark Reactions on Moodle.

18 November 2013: Tests will be returned to you during Wednesday's lecture. We will be continue to 'track the carbon' in the Calvin Cycle.

11 November 2013: Photosynthesis in the News Photosynthetic Machines

There will be a Public Lecture (Tuesday 26 November at 7:30 pm) by the 2012 NSERC Polanyi Prize Winner Gregory Scholes: Photosynthetic Machines: Why Nature is Astounding. He will be introduced by Nobel Laureate John C. Polanyi. "Photosynthetic solar energy conversion occurs on an immense scale across the earth, influencing our biosphere from climate to oceanic food webs. These are amazing solar cells!
Fronds in kelp forests, crustose coralline algae and purple bacteria show interesting properties that are helping reveal the chemical physics involved in the ultrafast energy transfers of light harvesting. This talk introduces the incredible processes that initiate photosynthesis in the first picoseconds after light is absorbed."

12 November 2013: Lab Exercises Here's the digest from Monday's lab. You should be able to compare the digest band sizes with the restriction map mounted at the Bio.Wiki.

10 November 2013: Term Test: Monday 18 November

4 November 2013: I mounted the ATP and NADPH production lectures on Moodle. We are now commencing the Dark Reactions of photosynthesis. The textbook provides good coverage in Chapter 7 (The Chemistry of Photosynthesis).

25 October 2013: Photosynthesis in the News Liquid Fuel

The New Yorker magazine published an article entitled Liquid Fuel, from the Sun, highlighting research coming from the Joint Center for Artificial Photosynthesis (JCAP). This is a heavily-funded research center whose mandate is "the development of an artificial solar-fuel generation technology". Most of the research is on the engineering side, but the biochemical-side of photosynthesis can provide some ideas, proven to work for 3500 million years!.

21 October 2013: I will be finishing up the lectures on Reaction Centers this week (I will update the lecture notes on Moodle when finished). The next lecture will be on ATP and NADPH production. I don't have a 'source' for this lecture. The textbook provides good coverage of both Electron and proton transport (Chapter 5) and Synthesis of ATP: photophosphorylation (Chapter 6). The lectures are relevant to the lab exercises you will be doing this week and next week (Labs 06 and 07), where you will be introduced to the Oxygen Electrode, a favorite tool of researchers studying chloroplasts (and mitochondria).

14 October 2013: Term Test: Wednesday 16 October

8 October 2013: Assignment I have updated my list of some examples of TIPS articles that might be suitable for your assignment [txt].

The lecture notes for Light Harvesting have been mounted on Moodle.


2 October 2013: Lab Exercises Here are before and after shots of chloroplast isolation from the green alga Eremosphaera viridis, courtesy of Knika's lab group. The highly birefringement chloroplasts in the lower panel indicate a high percentage of intact chloroplasts. To the right are leaves in situ. The chloroplasts are crescent-shaped (right panel).

2 October 2013: I've mounted the lecture notes for the fourth lecture topic (Pigments) on Moodle. The next lecture will be on Light Harvesting. I don't have a 'source' for this lecture. The textbook provides good coverage in Chapters three (Light Harvesting and Energy Capture) and four (Architecture of the Photosynthetic Apparatus).

26 September 2013: Photosynthesis in the News Oxygenic Photosynthesis gets Older

Canadian news sources have picked up on some recent research that has a national connection (UBC scientists). It's geological evidence for 'elevated' oxygen levels much earlier than previously thought (by 700 million years!). (CBC coverage). It's based on indirect metrics for oxygen levels (traces of oxidized chromium isotopes in old rocks) and predicts oxygen levels of about 0.002% (much lower than our present 20%). I think the finding will face scepticism. Like all evidence of early oxygenic photosynthesis, the data are frustratingly indirect. The sciency article was published in Nature.

24 September 2013: I've mounted the lecture notes for the second and third lecture topics (Bacterial Photosynthesis and Light) on Moodle. The next lecture will be on Photosynthetic Pigments. I don't have a 'source' for this lecture. The textbook provides good coverage of both Light (Chapter 2) and Light Harvesting (Chapter 3 --including pigments).

15 September 2013: I've mounted the lecture notes for the first lecture topic (Geological Photosynthesis) on Moodle. The next lecture will be on Bacterial Photosynthesis (anaerobic). The major objective is to introduce you to the diversity of photosynthetic mechanisms, highlighting alternative carbon dioxide fixation pathways. The long-term relevance is bioengineering. I don't have a 'source' for this lecture. A relatively recent overview is provided in an article by Bryant and Frigaard (2006) entitled Prokaryotic photosynthesis and phototrophy illuminated.

12 September 2013: Just a reminder for the labs on Monday and Tuesday. The reflectance spectroscopy uses fiber optic cables. Overbending the cables can damage the optical fibers. So handle them gently (don't force them to bend) during the labs.

10 September 2013: The first lecture topic (tomorrow) will be on Geological Photosynthesis. We will be exploring the different kinds of evidence supporting the appearence of photosynthesis in geological time. My source for this is a review by J. William Schopf (The paleobiological record of photosynthesis). His paper was part of a special issue of the journal Photosynthesis Research on the Evolution of Oxygenic Photosynthesis. An overview is available in your textbook (Chapter 1).

20 August 2013: Preparations for the Photosynthesis course are ongoing, including growing crops in the greenhouse to provide plants (and chloroplasts) for the lab exercises (online at Bio.Wiki). Students will select which kind of plants they want to use for their experiments. We've got beans, basil, sunflower, cucumber, corn and many other possibilities!

Update (14 September 2013): More recent snapshots of the plants to the right, the beans are already flowering!

13 August 2013: The textbook for Photosynthesis is available at the York Bookstore. The lab manual is available online at Bio.Wiki.

5 August 2013: Photosynthesis in the News Photosynthesis at the Microscale (CERL-30)

  • self assembled constructs that mimic one or more parts of photosystem II: photon capture, electron transfer, charge separation and water oxidation steps
  • membrane systems with selective passive transport of protons
  • immobilized hydrogenase mimic for reduction of protons to hydrogen or enzymes for CO2 conversion to biofuel.

22 July 2013: Photosynthesis in the News X-rays Illuminate Oxygen Evolution.

To see what happens during water splitting in photosystem II, you need to look fast and with extraordinary spatial resolution at physiologically relevant temperatures. That's where femtosecond x-ray pulses come into play: combining x-ray spectroscopy and x-ray diffraction. The work was done at the SLAC National Accelerator Laboratory in Menlo Park California. It revealed changes in the electronic structure of the Mn4CaO5 cluster that is central to the water-splitting reaction --light-driven oxidation that sequentially removes electrons (and hydrogen) from water, to eventually produce the oxygen byproduct. The sciency article was published in an April issue of Science Magazine. It's a good indication of how fast things are happening in the photosynthetic revolution.

25 June 2013: Photosynthesis in the News Algae Airships.

The BBC World News highlights futuristic visions of cities in a recent article, including an idea from the architect Vincent Callebaut: who proposed 'algae airships', using hydrogen-creating seaweed which allows the structures to float above the ground. There is a lot of interest in hydrogen-producing photosynthetic organisms. The idea of algal airships is a bit 'up in the air' (pun decidedly intended).

21 June 2013: Photosynthesis in the News Photosynthesis Goes Quantum.

18 April 2013: The tentative schedule for Photosynthesis labs is available (click on the image for a larger version). The lab manual is available online at Bio.Wiki.

9 April 2013: This will be the course website for the next offering of the course in the fall of 2013.

18 December 2012: Useful resources for the Photosynthesis course can be found at Hansatech Instruments.

18 December 2012: Photosynthesis in the News The Gates Foundation Sees the Light.

17 December 2012: The 2013 Course Outline is available in draft form [pdf].

28 December 2011: Lab Exercises The lab manual has been mounted as a LabWiki at the BioWiki website.

5 December 2011: In last week's lecture, I mentioned Black Leaves, an idea proposed by the futurist Freeman Dyson as a means to enhance the quantum efficiency of light capture in photosynthesis. Here are direct quotes [pdf] from Dyson's essay and a response from a Biochemist.

30 November 2011: I mounted the lecture notes for the ninth lecture (Metabolic Flux) on Moodle.

28 nov 2011: NSERC USRA Awards

28 November 2011: I mounted the lecture notes for the eighth lecture (Dark Reactions) on Moodle.

28 November 2011: The Final Exam is scheduled for Friday 16 December at 14:00 in CB (Chemistry Building) Room 115. The duration of the final is 3 hours. I expect the required time to be about 2 hours, but the additional time will be available to you as required.

15 November 2011: Assignment Reminder: Current Topic in Photosynthesis. As detailed below, please don't forget your upcoming assignment!

9 November 2011: As promised, a sample work problem on Gibbs free energy [png].

7 November 2011: I mounted the lecture notes for the seventh lecture (NADPH and ATP Production) on Moodle.

7 November 2011: Reminder: Second Term Test next Monday 14 November.

1 November 2011: I mounted the lecture notes for the sixth lecture (Reaction Centers) on Moodle.

27 October 2011: Lab Exercises Here is an example [png] of oxygen evolution in a whole cell suspension of Eremosphaera viridis that shows the effect of adding bicarbonate (that is, carbon dioxide for fixation). Note that dark causes oxygen consumption due to mitochondrial respiration.

25 October 2011: Assignment: Current Topic in Photosynthesis.

24 October 2011: Photosynthesis in the News The End Is Near.

An old news article reporting on a presentation at the American Association for the Advancement of Science highlights a long-standing concern of scientists, albeit with the excessive prose of the news media ("Combustion Leads to Eventual Suffocation"). Human consumption of resources, the burning of fossil fuels, the massive strain on global carrying capacity are real problems. The fact that The End Has Not Happened forty four years later may (or may not) be reassuring. It is all intertwined with photosynthesis.

20 October 2011: Photosynthesis in the News Aurora | algae Enters the Commercialization Phase.

20 October 2011: Photosynthesis in the News Press Releases.

19 October 2011: Photosynthesis in the News Practical Artificial Leaves.

19 October 2011: Photosynthesis in the News Global photosynthesis faster than thought.

18 October 2011: Lab Exercises Here are operating instructions for the Clark oxygen electrode [png] and chart recorder [png] (both will be provided as hardcopy for the oxygen evolution labs).

12 October 2011: Term Test: Monday 17 October

9 October 2011: Photosynthesis in the News Mapping Global Fluorescence.

7 October 2011: I mounted the lecture notes for the fifth lecture (Light Harvesting) on Moodle.

3 October 2011: I mounted the lecture notes for the fourth lecture (Pigments) on Moodle.

26 September 2011: As promised, a taste of quantum tunneling. It's new, fascinating, and a bit incomprehensible to your prof!

26 September 2011: I mounted the lecture notes for the third lecture (Light) on Moodle.

19 September 2011: Here is the Assignments and Grading decided on by the students:

15 September 2011: See you in lab this afternoon! (Room 106 Lumbers).

15 September 2011: I mounted the lecture notes for the second lecture (Bacterial Photosynthesis) on Moodle [link].

12 September 2011: I mounted the lecture notes for the first lecture (Evolutionary Photosynthesis) on Moodle [link].

11 September 2011: I mounted scans of overheads from the introductory lecture on Moodle [link].

7 September 2011: I hope you get the opportunity to discuss possible grading and assignment schemes amongst yourselves fairly soon. The University deadline for 'announcing components of final grades' is 20 September.

20 August 2011: Photosynthesis in the News Eating Sunshine.

15 August 2011:The moodle website for the course has been activated.

10 August 2011 update:The textbook (David W. Lawlor: Photosynthesis 3d edition) (and lab manual) are available at the York Bookstore [png]

19 June 2011: Photosynthesis in the News I thought students would be interested to know about the course they are taking from a broader perspective. It is not common to have a course on Photosynthesis at the undergraduate level, but there are Universities that do.

15 June 2011: Lab Exercises Here are operating instructions for the Clark oxygen electrode [png] and chart recorder [png] (both will be provided as hardcopy for the oxygen evolution labs).

3 June 2011: Here is a tentative schedule for lab exercises in the fall term (subject to change, as required) [png]

8 February 2011: This will be the course website for the next offering of the course in the fall of 2011.

Thank you all for making this such an enjoyable course, and, best wishes on all your future photosynthetic endeavours

Watch the video: Lecture 07, concept 19: Cytochrome C (August 2022).