Question

Q4. Compound A is labeled at the methyl carbon with 14C (*). Under fasting conditions, the 14C label ends up in compound C during steady-state metabolism in liver. COO- çoo- FO +H COO- H *CH3 a. What is the metabolic pathway involved? Write the reaction including all reactants and products, no structures required (1) b. Which carbons in compound C will become labeled, and to what percent? Briefly explain. (3) c. What is the physiological basis and the molecular mechanism for regulation of this reaction in response to glucagon? (3)

Answer 1

ANSWER 4.A :-

1. Steady-state basically means homeostatic measures which in case of chemical reactions within a cell deals with the reactions carried out by initial steps in the formation of the substrate and the intermediate reaction involving the substrate wherein it is converted in the presence of enzymatic activity into another product. So, this in turn is responsible for maintaining the sustainability of the reaction.
2. The compound C which is mentioned in the diagram is the Oxaloacetate which in turn is coupled with the modified form of Pyruvate called as Acetyl-CoA which is used for the entry of the substrate into the mitochondria.
3. During fasting, there is reduction of ATP molecules due to the TCA (Tricarboxylic acid cycle) followed by electron transfer chain which in turn leads to the synthesis of ATP molecules. Thus, there needs to be a replenishment of the (NADH + H+) which in turn is essential for the activation of the ATP synthetase activity mediated by the proton gradient set by Hydrogen ions.
4. There are a total of 8 reactions which are occuring in the entire process wherein the final product that is oxaloacetate is produced which again enters into the cycle.
5. During the first step, there is basically a reaction occuring in which the Pyruvate molecule is being converted into Acetyl-CoA molecule for entry into mitochondria where the first step is catalysed. This conversion is carried out by the enzyme complex of Pyruvate dehydrogenase. As stated above, oxaloacetate again enters into the cycle by means of condensation reaction being carried out wherein in the presence of water molecule there is linking of both the Acetyl-CoA and oxaloacetate leading to the formation of Citrate which is the first product of the reaction process. This reaction is catalysed by the enzyme Citrate synthase with the removal of Co-enzyme A molecule to form the complex substance. Citrate is basically a 6 carbon compound.
6. In the next step, there is a simultaneous dehydration and hydration reaction which is carried out wherein the enzyme cis-Aconitase leads to the formation of Isocitrate by enchanging the hydroxyl groups through the structure.
7. The next step is oxidative decarboxylation wherein there is removal of two hydrogen bonds and a carboxyl group in the form of CO2 leading to the formation of product called alpha-ketoglutarate. This reaction is catalysed by the enzyme isocitrate dehydrogenase. There is formation of one NADH + H+ during this step.
8. In the next step, there is conversion of the substrate into Succinyl-CoA which is carried out in the presence of the enzyme alpha-ketoglutarate dehydrogenase which is a complex associated with three catalytic molecules that bring about oxidative decarboxylation reaction including removal of carbon dioxide and formation of NADH + H+.
9. In the next step, the substrate Succinyl-CoA is converted into succinate in the presence of the enzyme Thiokinase wherein there is substitution of a phosphate group along with simultaneous removal of Co-enzyme A group and this is followed by substrate level phosphorylation wherein there is removal of phosphate which is accepted by the GDP molecule followed by simultaneous conversion of ADP to ATP mediated by phosphate transfer from GTP to ADP to form ATP.
10. In the next step, there is conversion of Succinate to Fumarate which is carried out in the presence of enzyme called as Succinate dehydrogenase associated with removal of hydrogen atoms and formation of a double bond between C2 and C3 carbon atoms. The hydrogen atoms are accepted by FAD to form FADH2.
11. This reaction is followed by the conversion of Fumarate to Malate in the presence of enzyme Fumarase. This reaction occurs with the addition of water molecules.
12. In the last step, there is conversion of Malate to Oxaloacetate usually carried out in the presence of enzyme Malate deehydrogenase which leads to the removal of hydrogen atoms which are being accepted by the NAD to form NADH + H+. This is the way in which the end product that is Oxaloacetate is produced which again enters into the Kreb’s cycle.

ANSWER 4.B :-

1. Before answering this question, the basic reaction where Pyruvate is converted to Acetyl-CoA needs to be considered. Every three carbon compound enters into the mechanism where there is removal of a carbon dioxide for each and subsequent oxidation process wherein there is removal of hydrogen carried out in the presence of the enzyme pyruvate dehydrogenase.
2. So, both are then linked to subsequent Co-enzyme A leading to the formation of Acetyl-CoA. The second carbon atom that is the one that is the keto form will be having the incorporation of C-14 radioisotope. Although regarding the amount of the substance being expressed can be traced by means of quantitative estimation.

ANSWER 4.C :-

1. Gdlucagon is the hormone which is involved in the upregulation of the mechanisms which increases blood glucose levels. This includes Gluconeogeneis, Glycogenolysis wherein glycogen is being cleaved in the presence of enzyme Glycogen phosphorylase which leads to the removal of Glucose-1-phosphate that can be subsequently converted Glucose-6-phosphate by means of phosphogucomutase enzyme activity.
2. During fasting state, the blood glucose levels falls down accompanied by sebsequent increase in the levels of FAD due to most of the FADH + H+ being used in the sythesis of ATP.
3. Once, the levels of glucose are back to normal, there will be activation of the Glycolysis pathway followed by Kreb's cycle to bring about enhanced synthesis of ATP. This is how Glucagon will be involved in positive response with respect to the low blood glucose levels.
4. Glucagon seretion will be stimulated depending upon blood glucose levels accompanied by increase in the number of receptors for the hormone to bind and mediate the activity. This will lead to binding of the hormone with the G-protein coupled Glucagon receptor followed by activation of Adenylate cyclase which triggers conversion of ATP to cAMP thereby leading to activation of Protein Kinase A activity depending upon the levels of cAMP. This will lead to phosphorylation and subsequent activation of Glycogen phosphorylase and other associated enzymes to bring about formation of glucose. As the levels of glucose comes back to normal, there will be activation of the Kreb's cycle to initiate ATP synthesis.