Question

M Low High Low Glycolysis CASALS 03. I am studying newly discovered cells in the human body. Within these cells, I find a novel protein, BFF. I purify BFF from these 50 KD - cells when an individual is 20 KD - fasting (Low), and after Low High Low Low High Low feeding (High), and run the purified BFF in a native PAGE. Using this native PAGE, I perform a Western blot, using a BFF-specific antibody. I also repeat these experiments with cells from patients that have a mutation in the BFF protein (in "High" conditions, the mutant behaves normally, results for "Low" conditions are marked LowM). My results for the Western are shown, along with measurements of Glycolysis and levels of CAMP in these cells. Levels of glucagon in the mutant are normal in all conditions. a. What is the MOST LIKELY pathway schematic that describes BFF function in cells? BFF → Adenylate cyclase CAMP Decreased glycolysis Glucagon → CAMP → Protein kinase A → BHF→ Decreased glycolysis N BFF → Glucagon → CAMP= Protein kinase A Decreased glycolysis Glucagon BFF → CAMP → Protein kinase A → Decreased glycolysis usis +PO4 b. What combination of data BEST supports your hypothesis for 3a? In BFF mutants, there is no effect on glucagon levels, but CAMP and glycolysis are affected in low glucose conditions only

im having a hard time undertsbading ghe rationale behind the asnwers (highlighted in pink). Please explain in detail an pathways/regukators that lead to the three answers, thank you!

Answer 1

ANSWER A :-

1. First of all, the hormone called as Glucagon plays an important role in increasing blood glucose levels. One important mechanism that occurs is inhibition of Glycolysis pathway as it is associated with increasing ATP concentration.
2. So, the ultimate effect can be seen in the form of inhibition of Glycolysis process but we need to identify which prior steps aids in the inhibitory response. It should be clear that in the absence of Glucagon hormone, the mechanism cannot be initatied as it binds to specific Glucagon G-protein coupled receptors to mediate their activity. This binding is accompanied with the activation of the enzyme Adenylate Cyclase which leads to increasing the cytosolic cAMP levels which in turn leads to phosphorylation events that will lead to stimulation of the glycogenolysis process releasing Glucose-1-phosphate molecule which can be converted to Glucose-6-phosphate after activity carried out the the enzyme Phosphoglucomutase.
3. It should be clear that the rate of Glycolysis will be low during fasting as there will be enhanced secretion of Glucagon which in turn will inhibit the process. At the same time, the levels of cAMP will be higher taking into fact the point mentioned above. In the mutant forms, exact opposite mechanism is seen which in turn supports the activity of enzyme insulin and hence indiates a regulatory control over glucose metabolism mediated by the protein BFF.
4. So, it should interact with the Glucagon hormone to mediate its activity although the mutated is stated to support the fact that insulin secretion would not be inhibited. This indicates that BFF protein might be associated with inhibition of insulin activity which is altered in the mutated form. So, prior to Glucagon hormone, it should be the BFF protein which should be mentioned taking into consideration its regulatory function. So, Option 'C' should be the most appropriate answer answer BFF binding would lead to activation of Glucagon mechanism after its inhibitory role mediated through insulin hormone. This is followed by the activation of adenylate cyclase which increases cytosolic cAMP levels and thus leads to activation of the Protein Kinase A thereby leading to activation of Glycogenolysis pathway.

ANSWER B :-

1. There is no appropriate data mentioned in the query above but the second and the third graph indicating rate of glycolysis and the levels of cAMP proves to be more effective in evaluating the results. I dont think the answer for A is the one indicated in pink because before activation of hormone Glucagon could not be initiated without inhibition of the hormone insulin. As can be seen in mutant form, it is low level of cAMP during fasting conditions where the levels should have been more and the high rate of glycolysis which prompts the correct answer to be option C.

ANSWER C and D -

1. The hormone Glucagon may lead to a signal which may be associated with activation of pathway for the degradation of the protein BFF although its not prominently seem to affect this as high glucose levels will directly prompt secretion of Insulin which in turn controls the secretion of the hormone Glucagon. So, there is no association with degradation of the protein.
2. cAMP is basically elevated only after activation of Adenylate cyclase activity but from the points mentioned above, there is no association of cAMP with the activation of BFF protein. Although, it should be taken into consideration that cAMP levels are low in mutated form but they should have been higher. This indicates that levels are in turn affected by the protein and they do not affect the protein directly. So, it should be the prior molecules or mechanism which should be mediated. Protein Kinase A is being activated depending upon the levels of cAMP and so its incuded in secondary signalling and there is no data indicating it affects the activity of BFF protein either positively or negatively and hence the option 'B' remains as the only preferable option mainly due to the fact that polymerization is associated with active nature and as blood glucose levels falls down, there is simultaneous increase in Glucagon hormone receptors and thus an associated activity would lead to polymerization and thereby activating the catalytic subunit of the protein. But, this may occur after its inhibitory role on insulin taking into consideration the levels of blood glucose.