alternative explanation for AAMC SB C/P #78?

[sapientnarwhal]

Experiment: A kinase (GalK) that phosphorylates a substrate (galactose) is coupled with pyruvate kinase and lactate dehydrogenase. Reaction is monitored based on decreasing amounts of NADH.
(table is given with info not relevant to question)

Question: Which molecule is expected to be in the lowest concentration at the completion of the reaction/end of kinetic assay?
a) lactate
b) ADP
c) ATP
d) NAD+

I knew that A and D were incorrect, as both are expected to accumulate. The correct answer is B, though I argue it could also be C. I believe both answers can be justified, depending on how one interprets the passage.

I reasoned that because the reaction is determined by decreasing NADH concentrations, the kinetic assay ends when most of the PEP has been converted to lactate. Based on this assumption, I reasoned that the "last" (in other words equilibrium is achieved) ATP molecule that pyruvate kinase produces is then subsequently hydrolyzed by the GalK to produce P-galactose and ADP.

Do I have a case here?

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[esob]

I missed this too but it is pretty straight forward when you read the 2nd sentence of the last paragraph. It tells you there are two reactions going on in the assay:
1. PEP to Pyruvate, assisted by Pyruvate kinase.
2. Pyruvate to Lactate.

When deciding between ADP or ATP, 1. yields ATP by adding moving the phosphate from PEP to ADP. Thus, that reaction should always end with more ATP than ADP, which means ADP concentration will be lowest.

 

[sapientnarwhal]

@esob
I understand that there are three reactions occurring. However, the reaction is monitored on the basis of decreasing NADH levels. This means that as the reaction proceeds to completion, lactate is increasing, NAD+ is increasing, Galactose-P is increasing, PEP is decreasing, and Pyruvate is decreasing.

From the limited passage information I think it is reasonable to argue that the reaction catalyzed by pyruvate kinase provides the ATP necessary for the GalK reaction.
- There must be a limited amount of PEP and Galactose in the reaction mixture, yet we do not know if the galactose substrate is consumed first or if the PEP is consumed first.

if the galactose is consumed first, then it is likely that the resultant ADP is phosphorylated by pyruvate kinase. Therefore ADP is in the lowest concentration.

if the PEP is consumed first, then it is likely that "all" of the ATP is hydrolyzed by GalK to phosphorylate galactose. Thus ATP would be least concentrated.

My contention is that both answers would work, neither are necessarily wrong. The passage did not provide any information that would contradict either conclusion.

I will take a look at this tomorrow to confirm that I'm not just tired.