Biochem Exam 3

A student set up an in vitro respiration system using cell extracts in which the rate of carbohydrate metabolism could be measured by monitoring the conversion of radioactive glucose to CO2. The student found that the addition of citrate to this system led to a rapid decrease in the rate of glucose metabolism; however, the addition of acetyl-CoA had little effect on the rate of glucose metabolism. Explain this observation.

Addition of citrate increased the capacity of the citrate cycle to metabolize acetyl-CoA by increasing the concentration of all cycle intermediates—particularly oxaloacetate. The addition of acetyl-CoA to this system had little effect because oxaloacetate levels were limiting. The cycle was working at full capacity, so having more substrate available had little effect on the rate of glucose metabolism.

The yeast Saccharomyces cerevisiae is used in the production of beer because of its ability to shift readily from aerobic respiration, in which it converts glucose to CO2 and H2O, to anaerobic respiration, in which it converts glucose to C2H6O (ethanol) and CO2. Explain why, in the production of beer, the yeast are first grown under aerobic conditions, and then shifted to anaerobic conditions. Why not just grow them under anaerobic conditions the entire time?

Aerobic conditions during the first phase of beer production facilitate rapid growth from the yeast starter culture to grow sufficient amounts of yeast (aerobic growth is much faster than growth under anaerobic conditions because of differences in efficiency of ATP energy generation). Then the beer mixture is shifted to anaerobic conditions to initiate the fermentation process, using the sugars from the plant materials.

In terms of photorespiration and ATP requirements, what explains the observation that crabgrass, which uses the C4 pathway of carbon fixation, has a growth advantage over turfgrass when temperatures are high and the O2-to-CO2 ratios are elevated because of increased O2 solubility?

Photorespiration is a "wasteful" reaction because 2-phosphoglycolate must be salvaged by the glycolate pathway at the expense of ATP hydrolysis. Therefore, the C4 plant has an advantage in summer because it minimizes loss of carbon to photorespiration when O2-to-CO2 ratios are elevated. However, the C4 pathway requires an investment of ATP to regenerate metabolites in a reaction that is not required by C3plants. Therefore, at low temperatures, when the CO2-to-O2 ratio is still relatively high, this extra ATP investment by C4 plants comes at an energy cost, thereby giving C3 plants a growth advantage.

The addition of 14C-pyruvate to a preparation of mitochondria leads instantaneously to 14CO2 production. However, adding 14CO2 to the mitochondria does not result in 14C-pyruvate. Why not?

Pyruvate is decarboxylated by the pyruvate dehydrogenase reaction to yield 14CO2 and acetyl-CoA. The reverse reaction is highly unfavorable and does not occur in the cell.

The key event in converting solar energy into redox energy is photooxidation of chlorophyll in the reaction centers. What must be the chemical property of the electron carrier molecule that accepts the electron from chlorophyll; that is, why is the electron transfer favorable?

The e− carrier molecule that accepts the e− from chlorophyll must have a higher reduction potential (the biochemical standard reduction potential is more positive than chlorophyll).

What redox reaction in PSII ultimately replaces the transferred electron so that the chlorophyll molecule becomes reduced and can therefore undergo another round of photooxidation?

The oxidation of H2O generates the e− that reduces chlorophyll.

Why does the ATP exchange ratio in mitochondria count 4 H⁺ translocated for every ATP synthesized, when only 3 H⁺ need to cross the inner mitochondrial membrane to generate 1 ATP?

The phosphate translocase requires that 1 H⁺ accompanies each Pi brought into the matrix.

What are the three metabolic fates of pyruvate? Include in your answer the conditions dictating which of these three metabolic fates is most likely to occur.

Under aerobic conditions, pyruvate is decarboxylated and converted to acetyl-CoA in organisms that use the citrate cycle. This yields the maximum number of ATP from glucose oxidation. Under anaerobic conditions, pyruvate is converted to lactate by lactate dehydrogenase in order to regenerate NAD⁺ for the glyceraldehyde-3-phosphate dehydrogenase reaction in glycolysis. Alternatively, fermenting organisms, such as yeast, convert pyruvate to CO2 and ethanol under anaerobic conditions.

Compare and contrast the functions of the malate-aspartate and glycerol-3-phosphate mitochondrial shuttle systems.

Both shuttles serve to donate electrons from mitochondrial NADH into the cytosol to regenerate NAD⁺ for the glycolytic pathway. The malate-aspartate shuttle in liver cells uses NADH to reduce cytosolic oxaloacetate and generate NAD⁺ and malate, which is shuttled into the matrix and oxidized to produce NADH and oxaloacetate. The glycerol-3-P shuttle in muscle and brain cells catalyzes a redox reaction that oxidizes cytosolic NADH and reduces FAD to yield cytosolic NAD⁺ and mitochondrial FADH2. Oxidation of the FADH2 within the inner mitochondrial membrane transfers 2 e− directly to coenzyme Q . The malate-aspartate shuttle in liver cells converts 1 NADHcytosol to 1 NADHmatrix, the maximum yield of redox energy. In contrast, the glycerol-3-P shuttle in muscle and brain cells provides less ATP (oxidation of FADH2 in the electron transport system results in the translocation of fewer H⁺ across the inner mitochondrial membrane).

Carbon dioxide is a C1 substrate in the reaction of the Calvin-Benson cycle that is catalyzed by the enzyme RuBisCO. What are the one C5 substrate and two C3products of this reaction? Why is the mass of RuBisCO on Earth higher than that of any other enzyme?

C5: ribulose-1,5-bisphosphate; C3: 3-phosphoglycerate. The abundance of RuBisCO reflects the importance of carbon fixation and that it catalyzes this reaction slowly. It is found in plants as well as algae and cyanobacteria.

An ATP synthase complex has been characterized that contains a c ring with 12 identical c subunits. Experiments have shown that for this ATP synthase, 1 ATP is synthesized for every 4 H⁺ that move through the complex into the matrix. Explain this 1:4 ratio in terms of the structure and function of the γ and β subunits of the ATP synthase complex.

When the H⁺ binds to a single c subunit, it causes a rotation of the c ring of 30° (1/12 of a circle). The three-sided γ subunit rotates along with the c ring and interacts directly with the three β subunits, changing from L, T, or O conformation for every 120° rotation of the c ring. Because an ATP is synthesized for every turn of 120°, 4 H⁺ must cross through the ATP synthase complex to synthesize 1 ATP.