[latent_lamp]

Well, we did a simple oxygen electron experiment (mentioned in Voet n Voet) to measure the effectiveness of certain inhibitors on patato mitochondria. The experiment sucked, primarly because I think our mitochondria went into strike in the controlled phase (with succinate and ADP) so no surpises when KCN was added that it didnt do much.

But one of the questions posited by our beloved and estranged lecturer was that an excess of EITHER pyruvate OR malate stops oxygen consumption in mitochondria and when the other is added (equal amount) then o2 consumption resumes.

first I went WTH, not being the studious fellow I am, I took at a stab in the dark. Considering mitochondria are amphibolic; it is highly likely that the catabolic processes are inhibited and anabolic initiated when there is an excess of either of the two substrates (assuming that the enzymes involved have some form of regulation (i.e. phosphorylation)). I had heard that an excess of malate triggers lactate production in mammals (yes, highly likely that a patato with lactate) and also that within the pancreas the equilibrium of Acetyl-COA is maintained by malate/pyruvate shuttle.

Those explanations dont really explain what is going on in mitochondria of patato. I am not certain but confident that there is an inversion of metabolic pathways but to what? What is malate and or pyruvate converted to in a patato if my explanation stands.

I've already given the report in with my stupid explanations, so yeh this is just for self satisfaction.

[Equi]

Consider the TCA cycle: Without acetyl-CoA the cycle wouldn't run. Therefore pyruvate is essential to get the cycle running. High levels of pyruvate and low levels of intermediates lead to acetyl-CoA accumulation and inhibition of PDH and other key enzymes.

High levels of malate without acetly-CoA - well there's nothing to utilise.

However, when ATP accumulates the whole thing stops anyway. Did you use uncouplers?

[latent_lamp]

 Many thanks for your reply.


Essentially the mitochondrial TCA and ETS were coupled, we didnt do anything to cause them to divorce each other

The little understanding I have of TCA, is that pyruvate is one of those substrates that regulate gluconeogenesis relative to glycolysis. Malate supposedly does a similar thing (in mammals, malate is what is converted into lactose). Now that you mention Acetyl-COA, I must add that we didnt do anything that might inhibit any enzyme or cause diminution (by comsumption) of prexisting substrates.

So all the other stuff was already there (i.e. a-ketoglutarate, isocitrate and the rest), what we did was increase the concentration of Pyruvate or Malate and we found that 02 consumption completely stopped. But when the other was added then it would continue.

Now that particular observation is enough to say that both are in equilibrium of one another and therefore disequilibrium of these two substrates affects the oxidative catalysis of glucose.

The batch solution (for controls, inhibitors (we also tested inhibitors, but different batches), and this particular experiment) was the same, that being it had ADP, Succinate, and a phosphate buffer.


Big question is why??

[Equi]

The batch solution (for controls, inhibitors (we also tested inhibitors, but different batches), and this particular experiment) was the same, that being it had ADP, Succinate, and a phosphate buffer.


Big question is why??

I'm not quite sure to what your "why" refers. I hope I'll answer the right part

Since the citric acid cycle is a anaplerotic pathway it can generate the other intermediates out of a single one. Succinate is past the decarboxylation steps and hence every intermediate up to oxaloacetate is produced, the others when pyruvate is added and acetyl-CoA enters the cycle.

Phosphate serves an important shutteling role in the complex transport mechanism of mitochondria (most are electroneutral). Besides being part of the buffer and used in oxidative phosphorylation.

The control of glycolysis and gluconeogenesis is a bit more complex. Citrate plays a central role in inhibiting phosphfructokinase.

[latent_lamp]

Well the essay is already handed in (a long long time ago), I will go bug my proffessor and provide the answer that he expected...from the horses's mouth as it were.
I spoke to my tutor for this unit (second year intro biochem), since he is a Dr in microbiology, he basically said such things are useless to him (i.e. not his field of research), little help for me. That was the reason why I actually even asked on the forums.

I had exhausted all my resources, logic, textbooks, journals and even tutors without avail. I had found many answers to this question but none pertaining to patato mitochondria. I mean it would be stupid of me to write about a lactose pathway when I am sure patatoes dont lactate....well not the ones I eat...lol