Definitions:
End-product inhibition is the process whereby the last compound in a metabolic pathway acts as an inhibitor of the enzyme catalyzing the first committed step in the pathway leading to that compound. An example is the inhibition of aspartate transcarbamoylase by CTP.
[ 2 points for a reasonable definition, 2 for a plausible example; this one is not the only one].
A steroid hormone is a cholesterol derivative that acts as a hormone, i.e. is a circulating molecule that binds to cellular receptors to signal changes in metabolism. Testosterone, aldosterone, estrogen, and progesterone are examples.
[ 2 points for the definition, 2 for the example]
Gluconeogenesis is the ATP-dependent pathway by which pyruvate molecules are used as starting molecules to build up toward production of new sugar phosphates and thence to glycogen.
[ 2 point for the fact that it is a pathway toward sugars; 1 for recognizing that the starting metabolite is pyruvate; 1 point for stating that the pathway requires energy or that it requires expenditure of ATP; For students who would otherwise receive less then full credit, give 1 bonus point for recognizing that the end products are sugar phosphates and / or glycogen, not glucose itself.].
A ketone body is a metabolite containing a keto or sec-alcohol group, derived from lipid catabolism via acetyl coenzyme A. Acetone, acetoacetate, and β-hydroxybutyrate are examples.
[2 points for a reasonable definition; 2 for one of those three examples. If the student named all three examples correctly then give full credit no matter what else is there.]
Short-Answer Questions:
[ 2 points for glycolysis; 2 points for carbon-carbon.]
[2 points for dihydroxyacetone phosphate; 1 point for barrel.]
[ 2 points for 8 Acetyl CoA molecules; 2 points for NADH. If someone puts down QH2 or reduced quinone instead of NADH, give full credit. If someone puts down H+ or H3O+ instead of NADH, give one point for the second half.]
[one point for each blank. I will allow "relaxed" as a synonym for "loose" and "tense" or "tensed" as a synonym for "tight".]
[ Two points for each answer. No credit for abbreviations, since I explicitly said to avoid using them. But "high-density" for the first answer (without "lipoproteins") will earn one point.]
Calculational problems:
Palmitate has formula CH3(CH2) 14COOH, or C16H32O2, so its molecular weight is 16*12+32+32= 256.
Thus 1 g of palmitate is(1 g / 256 g/mol) = 3.91*10-3 mol.
4.184 J = 1 cal, so 4184 J = 1 kcal.
Overall energy produced =
(3.91 * 10-3 mol palmitate) * (106 moles ATP per mole of palmitate) *
(7.3 kcal /mol ATP) * 4184 J /kcal) = (3.91 * 1.06 * 7.3 * 4.184) * 10-3+2+3 =
126.6 * 102 = 126 * 100 = 12600 J.
[ 3 points for getting the basic algebraic approach right. 2 points for getting the molecular weight of palmitic acid correct and using it right. 1 point for getting the conversion from kcal to J right. 2 points for the right number to within 5%. If the answer is left in kcal but is the correct number of kcal (3.01), award up to 7 points. (Following instructions does matter)!]
(a) Compute the total protonmotive force Δp associated with moving one proton across such a membrane at 25°C. Note that Faraday's constant F = 96.485 kJ V-1 mol-1 and the gas constant R = 0.001986 kcal mol-1deg-1 = 0.008314 kJ mol-1deg-1. (4 pts)
As Horton works out in detail on p. 439, Δp = ΔG/(nF) = Δψ - 2.303 RTΔpH / F
At 25°C = 298K, 2.303 RT / F = 2.303 * 0.008314 * 298 / 96.485 = 0.05913 V,
so (Horton, p.439, eqn. 14.15), Δp = Δψ - (0.059 V) * ΔpH
Thus for Δ&psi = -0.017 V, ΔpH = 0.5, Δp = -0.17 V - (-0.59 V) * (0.5) = -0.20 V.
[ 2 points for the correct equation and approach; 1 point for getting the algebra correct; 1 point for the correct answer to within 5%. ]
(b) What is the total free energy obtained from this transaction? (4 pts)
We know that ΔG = nF Δp, so to pass one electron through the membrane, n = 1, so ΔG = 1 * (96.485 kJ V-1mol-1) * (-0.20 V) = -19 kJ mol-1
[ 2 points for the correct formula for ΔG. 2 points for getting the answer right to within 5%. If the answer in (a) is wrong but the wrong answer is correctly carried through into (b), award up to 3 points. The answer can be expressed as -79 kcal mol-1 for full credit.]
Paragraph questions:
Answer three of the following five questions. Remember that we want a coherent paragraph, not four paragraphs, for each answer.Cholesterol acts as a component in phosphipid-bilayer membranes, modifying their bulk and local properties. It functions as a carrier for other lipid molecules within lipoproteins, including chylomicrons, VLDLs, IDLs, LDLs, and high-density lipoproteins. It is a metabolic precursor to the steroid hormones. Finally, its breakdown products, the bile salts, play metabolic roles in liver function and energetics. The structure is as follows:
[2 points each for the three roles; if all four are specified, you don't get any extra points. 2 points for getting the structure right.]
is broken down into smaller building blocks. Indicate where within the cell this degradation occurs and the benefit to the cell that accrues from this process. Is the double bond in oleate cis or trans? (8 pts)
Oleate is activated by conjugation to coenzyme A and broken down for the most part by beta-oxidation, through which two carbons at a time are removed from the backbone to yield molecules of acetyl CoA. In the vicinity of the double bond in the center of the molecule, some special reactions come into play wherein the trans double bond two carbons away from the carboxylate is isomerized to a cis double bond only one carbon away (i.e., conjugated to the carboxylate carbonyl) through the action of the enzyme enoyl-CoA isomerase). This is again a substrate for one of the enzymes involved in ordinary beta-oxidation, so the process proceeds normally after that.
This reaction occurs in the mitochondrion and it acts as a source of metabolic energy in the form of the ATP that can be produced from the product, acetyl CoA. It can also serve as a carbon source for the buildup of subsequent molecules of other fats. The double bond in oleate is cis.
[ 3 points for the breakdown pathway; 1 point for knowing that it happens in the mitochondrion; 1 point each for knowing that it's an energy source and for knowing that the product can be used to rebuild fats. 2 points for knowing that the double bond is cis. The details of the breakdown pathway need not be specified at the level of naming the enzymes.]
In both instances the protonmotive force is carried out through the actions of a multi-component ATPase enzyme system; the ATPases in chloroplasts and the ATPases in mitochondria are almost identical chemically, so they function almost identically. The protonmotive force in chloroplasts differs from that in mitochondria in that (1) in chloroplasts, there is passage of alkalai ions back across the membrane in opposition to the movement of protons, so the energy derived from the differences is almost purely due to chemical potential, whereas in mitochondria it is due to chemical and electrical potential; and (2) in chloroplasts the purpose of the energy is to build up metabolites in the dark reactions of photosynthesis, whereas in mitochondria the energy is used to build up catabolic metabolites of general use to the cell.
[ 2 points for identifying the ATPase as a highly similar component; 1 for understanding the a difference comes from how the energy derived is used; 1 for knowing that the electrical gradient is present in one case and not the other.]
(b) How does succinate-ubiquinone oxidoreductase differ from the other three primary electron-transport complexes in terms of its ability to contribute to the proton concentration gradient across the membrane? (4 pts)
Each of the other complexes (complexes I, III, and IV) embodies a sufficiently large electrochemical potential difference that it is capable of directly contributing to the proton concentration gradient across the inner mitochondrial membrane. The succinate-ubiquinone oxidoreductase system (complex II) has a ΔGo' of only -2 kJ/mol, so it does not contribute significantly to the gradient.
[ 2 points for knowing that it has a small ΔGo' value; 1 point for explicitly contrasting that small value with the larger ΔGo' value for the other complexes; 1 point for stating that the utility of this difference in free energy is to drive protons across the membrane.]
The two reactions are the carboxylation reaction and the oxygenation reaction:
[carboxylation:] ribulose 1,5-bisphosphate + CO2 → 2 3-phosphoglycerate
[oxygenation:] ribulose 1,5-bisphosphate + O2 → 3-phosphoglycerate + 2-phosphoglycolate
The significance to plant growth is that the carboxylation explicitly incorporates ("fixes") inorganic carbon (in the form of CO2 or HCO3-) into organic molecules, resulting in net opportunities for increasing biomass, i.e. allowing for plant growth. The oxygenation reaction, which is the primary step in photorespiration, contributes nothing to net carbon fixation, and in fact expends energy without incorporating new carbon into biomass. Therefore the amount of carboxylation that occurs in a plant, both absolutely and relative to the amount of oxygenation, is an important determinant of how rapidly a plant can grow.
[ 4 points for naming the reaction; 4 points for explaining the significance of the reactions to plant growth. In the reaction portion, drawing the structures correctly can substitute for naming the compounds. If the reaction is described correctly without naming the products explicitly, count off at most two points. In the explanation portion, be reasonably lenient about the description, provided that the concepts are there.
3-hydroxy-3-methylglutaryl Coenzyme A (HMG CoA), is a six-carbon precursor both of ketone bodies and of steroids. It is derived from acetyl CoA and acetoacetyl CoA, which in turn is derived from fusion of two molecules of acetyl CoA. Thus HMG CoA requires three molecules of acetyl CoA as starting materials. The path toward ketone bodies occurs through HMG-CoA lyase, which catalyzes the cleavage of HMG-CoA to acetyl CoA and acetoacetate, whence the other two common ketone bodies (acetone and β-hydroxybutyrate) can be derived--enzymatically in the case of β-hydroxybutyrate, and nonenzymatically in the case of acetone.. The path toward steroids occurs through HMG-CoA reductase, which catalyzes the reductive elimination of HS-CoA from HMG CoA to mevalonate. Mevalonate can be phosphorylated to phosphomevalonate and further phosphorylated to pyrophosphoromevalonate; this is then decarboxylated to produce isopentenyl pyrophosphate, an important 5-carbon intermediate. Fusion of isopentenyl pyrophosphate with an isomer, dimethylallyl pyrophosphate, gives rise to a C10 intermediate, geranyl pyrophosphate, which then fuses head-to-head with another molecule of isopentenyl pyrophosphate to produce a C15 intermediate, farnesyl pyrophosphate. Two molecules of farnesyl pyrophosphate fuse head-to-head to form the C30 intermediate squalene, from which the ring-form steroids, including cholesterol, are derived. The step catalyzed by HMGCoA reductase is the first committed step in the pathway toward steroid biosynthesis, so it is an important control step and has been an important drug target.
[No more than eight points can be earned, but the following portions of the answer will earn points: 2 points for recognizing that HMG CoA is a precursor of both ketone bodies and steroids; this is implied by the question, but the student should say it. 1 point each for naming a specific ketone body and a specific steroid or sterol. 1 point each for mentioning the names of the two enzymes that process HMG CoA toward a product. 1 point for describing the pathways that interconvert the ketone bodies. 1 point for describing the pathway from HMG CoA to isopentenyl pyrophosphate, apart from naming the first enzyme. 1 point for describing the path from isopentenyl pyrophosphate to farnesyl pyrophosphate, and one more for describing the path from farnesyl pyrophosphate to squalene and the C30 and C28 steroids. 1 point for mentioning that HMG CoA is a control point in the synthesis of steroids, and 1 point for mentioning that it is a drug target.]