BCSE Veterinary Biochemistry and Molecular Biology — Questions and Answers
Question 1: A cat with hepatic lipidosis has severely elevated plasma triglycerides. Which key enzyme in the TCA cycle is inhibited by the accumulation of NADH during excessive fat oxidation, perpetuating the cycle of hepatic fat accumulation?
- Pyruvate carboxylase
- Isocitrate dehydrogenase (Correct answer)
- Succinate dehydrogenase
- Aconitase
Correct answer: Isocitrate dehydrogenase
Isocitrate dehydrogenase is allosterically inhibited by high NADH/NAD+ ratios. During excessive β-oxidation (as occurs in feline hepatic lipidosis), NADH accumulates, inhibiting this enzyme and slowing the TCA cycle. Acetyl-CoA then cannot enter the cycle efficiently and is diverted toward ketone body and fatty acid synthesis, worsening fat accumulation. This NADH feedback inhibition is a central mechanism.
Question 2: Which base-pairing rule is violated in RNA but not DNA?
- Adenine pairs with thymine
- Guanine pairs with cytosine
- Adenine pairs with uracil instead of thymine (Correct answer)
- Cytosine pairs with guanine via three hydrogen bonds
Correct answer: Adenine pairs with uracil instead of thymine
In RNA, uracil replaces thymine. Thymine pairs with adenine in DNA (via two hydrogen bonds). RNA uses uracil instead, which also pairs with adenine but lacks the 5-methyl group of thymine. Guanine–cytosine pairing (three hydrogen bonds) occurs in both DNA and RNA. This distinction is fundamental to understanding transcription and RNA secondary structures.
Question 3: The Km of an enzyme refers to:
- The maximum reaction velocity when all enzyme active sites are saturated
- The substrate concentration at which reaction velocity equals ½ Vmax (Correct answer)
- The rate constant for product release
- The equilibrium constant for enzyme-substrate binding
Correct answer: The substrate concentration at which reaction velocity equals ½ Vmax
Km (Michaelis constant) is the substrate concentration at which the reaction velocity is half of Vmax. A low Km indicates high affinity (the enzyme reaches half-maximal velocity at low substrate concentrations). Vmax is the maximum velocity at enzyme saturation. Km is not the equilibrium dissociation constant (Kd), though it approximates it when the product release step is rate-limiting.
Question 4: In the polymerase chain reaction (PCR), what is the purpose of the denaturation step?
- To allow primers to anneal to single-stranded template
- To separate the double-stranded DNA template into single strands (Correct answer)
- To extend the new DNA strand from the 3' end of each primer
- To activate Taq polymerase by heat
Correct answer: To separate the double-stranded DNA template into single strands
Denaturation (typically 94–98°C) breaks the hydrogen bonds between complementary DNA strands, converting double-stranded DNA to single-stranded templates. This must precede primer annealing. Primer annealing occurs in the next step (lower temperature). Extension by Taq polymerase occurs at ~72°C. Taq is thermostable and does not require activation by heat.
Question 5: Ammonia toxicity in ruminants fed high non-protein nitrogen (NPN) diets occurs because excess ammonia overwhelms the liver's ability to convert it to urea via the urea cycle. Which enzyme catalyzes the rate-limiting, first committed step of the urea cycle?
- Arginase
- Ornithine transcarbamylase
- Carbamoyl phosphate synthetase I (CPS I) (Correct answer)
- Argininosuccinate lyase
Correct answer: Carbamoyl phosphate synthetase I (CPS I)
Carbamoyl phosphate synthetase I (CPS I), located in the mitochondrial matrix, catalyzes the ATP-dependent conversion of ammonia + CO2 → carbamoyl phosphate. This is the rate-limiting step of the hepatic urea cycle and is allosterically activated by N-acetylglutamate. When ammonia influx exceeds CPS I capacity (as with rumen NPN overload), blood ammonia rises, causing neurological signs.
Question 6: During oxidative phosphorylation, uncoupling agents such as 2,4-dinitrophenol (DNP) cause hyperthermia by:
- Directly inhibiting ATP synthase, halting ATP production
- Dissipating the proton gradient across the inner mitochondrial membrane as heat without synthesizing ATP (Correct answer)
- Blocking electron transfer at Complex III
- Inhibiting substrate-level phosphorylation in the TCA cycle
Correct answer: Dissipating the proton gradient across the inner mitochondrial membrane as heat without synthesizing ATP
Uncouplers are lipophilic weak acids that shuttle protons across the inner mitochondrial membrane, bypassing ATP synthase. The energy stored in the proton-motive force is released as heat rather than captured as ATP. Electron transport continues (and may accelerate), but ATP synthesis is uncoupled. This explains hyperthermia and cellular energy depletion seen in DNP poisoning and in conditions involving endogenous uncoupling proteins (thermogenesis in brown adipose tissue).
BCSE Veterinary Biochemistry and Molecular Biology
A cat with hepatic lipidosis has severely elevated plasma triglycerides.
Which key enzyme in the TCA cycle is inhibited by the accumulation of NADH during excessive fat oxidation, perpetuating the cycle of hepatic fat accumulation?