Problem Set on Enzyme Kinetics Phosphofructokinase (PFK) is a major control point in regulating the flux of metabolites into the glycolysis. A Biochemist has successfully isolated a potent specific inhibitor of PFK (inhibitor A) from an endemic plant in the Philippines. The following data describe the effect of the purified inhibitor on the activity of PFK. (20 points) Without Inhibitor With Inhibitor A [S] (mM) (μM⚫s¯¹) (μM⚫s¯¹) 1 2.5 1.17 10 2502 4.0 2.10 6.3 4.00 7.6 5.7 9.0 7.2 A. Construct a Michaelis-Menten and Lineweaver-Burke curves for the kinetic data above in the absence and presence of inhibitor A. B. Determine the Km, Km.app. Vmax, Xmax.app from the kinetic data. Show your complete calculation and encircle your final answers. Determine the mode of inhibition exhibited by inhibitor A Discuss your answer briefly. C. If the total PFK concentration is 1 nmol/L, how many molecules of substrate can a molecule of PFK process in each minute in the absence and presence of inhibitor A. Show your complete calculation and encircle your final answers. D. Determine the catalytic efficiency of PFK (in the absence and presence of inhibitor A. Is PFK a fairly efficient enzyme? Show your complete calculation and encircle your final answers.

Distinguishing the Mechanisms of Class I and Class I Aldolases Fructose bisphosphate aldolase in animal muscle is a class 1 aldolase, which forms a Schiff base intermediate between substrate (for example. fructose-1, 6-bisphosphate or dihydroxyacetone phosphate) and a lysine at the active site (see Figure I8.12). The chemical evidence for this intermediate conies from studies with aldolase and the reducing agent sodium borohydride, NaBH4. Incubation of the enzyme with dihydroxyacetone phosphate and NaBH4 inactivates the enzyme. Interestingly, no inactivation is observed if NabH4 is added to the enzyme in the absence of substrate. Write a mechanism that explains these observations and provides evidence for the formation of a Schiff base intermediate in the aldolase reaction.

Using the ActiveModel for aldose reductase, describe the structure of the TIM barrel motif and the structure and location of the active site.

Regulation of Glutamine Synthetase by Covalent Modification Suppose at certain specific metabolite concentrations in vivo the cyclic cascade regulating E. coli glutamine synthetase has reached a dynamic equilibrium where the average state of GS adenylylation is poised atn=6. Predict what change in nwill occur if: [ ATP ] increases, PIIA/PIID increases, [ -KG ]/[ Gln ] increases, [ Pi ] decreases.