2) In the early 1930's Albert Szent Gyorgyi reported the interesting observation that the addition of small amounts of oxaloacetate or malate to suspensions of minced pigeon breast stimulated oxygen consumption in of the preparation. Surprisingly, the amount of oxygen consumed was 7 times higher than the amount required for complete oxidation (to CO2 and H2O) of the added oxaloacteate or malate.
a) Why did the addition of oxaloactetae or malate stimulate oxygen consumption?
b) Why was the amount of oxygen consumed so much greater than the amount required to completely oxidize the added oxaloacteate and malate?
3) In the last reaction of the citric acid cycle, malate is
dehydrogenated to regenerate the oxaloacetate necessary for the entry of
acetyl coA into the cycle.
L-Malate + NAD+ -> oxaloacetate + NADH + H+ DGo'=30.5 kJ/mola) Calculate the equilibrium constant for this reaction at 25oC.b) Because DGo' assumes standard standard pH of 7, the equilibrium constant calculated is (a) corresponds to
Keq'= [oxaloacetate] [NADH]
[L-malate] [NAD+]
The measured concentration of L-malate in rat liver mitochondria is about 0.20 mM when [NAD+]/[NADH] is 10. Calculate the concentration of oxaloacteate at pH 7 in mitochondria.c) To appreciate the magnitude of the mitochondrial oxaloacetate concentration, calculate the number of oxaloacetate molecules in a single rat mitochodrion. Assume the mitochondrion is a sphere of diameter 2.0 mm.
4) Cellular respiration can be studied in isolated
mitochondria by measuring the oxygen consumption under different conditions.
If 0.01M sodium malonate is added to actively respiring mitochondria that
are using pyruvate as a fuel source, respiration soon stops and a metabolic
intermediate accumulates.
a) What is the structure of the intermediate that accumulates?
b) Explain why it accumulates.
c) Explain why oxygen consumption stops.
d) Aside from the removal of malonate, how might this inhibition of respiration be overcome? Explain.
5) An actively respiring bacterial culture is briefly incubated
with [1-14C] glucose. and glycolytic and citric acid cycle
intermediates were isolated. Where is the 14C is each of the following
intermediates listed below? Consider only the initial incorporation
of 14C, in the first pass of the labeled glucose through the pathways.
a) Fructose 1,6 bisphosphate.6) People with Beriberi, a disease cause by a thiamin deficiency, have elevated levels of blood pyruvate and a-ketoglutarate, especially after consuming a meal rich in glucose. How are these effects related to a deficiency in thiamin?
b) Glyceraldehyde 3-phosphate.
c) Phosphoenolpyruvate.
d) acetyl CoA
e) Citrate
f) a-ketoglutarate
g) oxaloacetate
7) Fluoroacetate, prepared commercially for rat control, is also produced by a South African plant. After entering the cell, fluoroacetate is converted to fluoroacetyl-CoA in a reaction catalyzed by the enzyme acetate thiokinase:
OThe toxic effect of fluoroacetate was studied in an experiment using intact isolated rat heart. After the heart was per fused with 0.22 mM fluoroacetate, the measured rate of glucose uptake and glycolysis decreased, and glucose 6-phosphate and fructose 6-phosphate accumulated. Examination of the citric acid cycle intermediates revealed that their concentrations were below normal, except for citrate, with a concentration 10 times higher than normal.
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F-CH2COO- + CoA +ATP -> F-CH2C-CoA AMP + PPi
a) Where did the block in the in the citric acid cycle occur? What caused citrate to accumulate and the other cycle intermediates to be depleted?b) Fluoroacetyl CoA is enzymatically transformed in the citric acid cycle. What is the structure of the end product of this metabolism? Why does this product block the citric acid cycle? How might this inhibition be overcome?
c) In the heart perfusion experiments why did glucose uptake and glycolysis decrease? Why did the hexose monophosphates accumulate?
d) Why is fluoroacetic acid poisoning fatal?
8) A deficiency of a citric acid cycle enzyme in both mitochondria
and the cytosol of some tissues (e.g. blood lymphocytes) results in severe
neurological abnormalities in newborns. The disease is characterized
by excretion in the urine of abnormally high amounts of a-ketoglutarate,
succinate, and fumarate. What enzyme deficiency would lead to these
symptoms?
9) Calculate the number of ATP molecules generated by the net reactions of the citric acid cycle for each of the reactions below. Assume all NADH2 and QH2 are oxidized to yield ATP, pyruvate is converted to acetyl CoA, and the malate aspartate shuttle is operating.
a) 1 pyruvate -> 3CO2
b) Citrate -> oxaloacetate
10) When one molecule of glucose is completely oxidized to 6 molecules
of CO2, under the conditions in Problem 9, what
percentage of ATP is produced by oxidative phosphorylation and what percentage
of ATP is produced by substrate level phosphorylation?
Problem #1