There are 10 problems in the problem set 3. Each problem will be graded on a 10 point scale, for atotal of maximum 100 points.For full credit, all steps to the solutions of the following problems must be shown. You may worktogether on the problems, but you may not copy or plagiarize. Your answers must be written, mustshow your solution steps and/or formulas when requested, and must be in your own words.Problem 1Glycogen phosphorylase converts glycogen into glucose subunits and is the rate limiting step inglycogenolysis (that will be treated later in the course). It catalyzes the following reaction:(α-1,4 glycogen chain)n + Pi ? (α-1,4 glycogen chain)n-1 + α-D-glucose-1-phosphateThe plot below indicates the rate of the reaction for glycogen phosphorylase as function ofphosphate concentration:a) Explain whether glycogen phosphorylase follows classic Michaelis-Menten kinetics ornot? Explain in detail your conclusion.b) Based on the plot, do you expect the enzyme to be a monomer or a dimer (note that asingle glycogen phosphorylase chain has a single Pi substrate binding site)?c) Caffeine is an allosteric inhibitor and glycogen phosphorylase is a K-system. How is therate curve affected and why?d) Plot the rate versus substrate curve for the enzyme in the presence of 1) ATP 2) Glucose3) AMP, knowing that the first 2 are feedback inhibitors while AMP is a positive effector.e) Draw the plot of the v versus substrate concentration for potential effectors in V-systems, and explain the rationale behind their effect on Vmax and KM (or K0.5). Problem 2Repeated vomiting and severe dehydration cause alkalosis, a condition in which serum pH ishigher than normal (7.45 or higher). Contrarily acidosis, occurring for example in patients withrenal failure, occurs when the serum pH fall below 7.35.a) Plot the curves of hemoglobin oxygen saturation for patients in conditions of alkalosis oracidosis, and compare it to the one in normal conditions.b) Explain why alkalosis can result in hypoxia even with sufficient O2 in the blood?c) In acidosis is oxygen released more or less efficiently to the tissues? What is the effect ofacidosis on hemoglobin in the lungs?d) Fetal hemoglobin possess a higher affinity for O2 by possessing two α and two γ subunitsinstead of two α and two β subunits. Explain why fetal hemoglobin shows an exacerbatedBohr Effect compared to adult hemoglobin.Problem 31) Draw and name the following monosaccharidesa) Anomer of β-D-galactoseb) Enantiomer of D-galactosec) The pyranose and furanose forms of D-galactosed) The epimer of D-galactosee) One of β-D-galactose diasteroisomerf) the corresponding aldopentose of D-talose with the same stereochemistry at each chiral(C-1) center2) Determine the relationship between the following pairs of sugars (anomers, enantioners,epimers, diasteroisomers, aldose and ketose pair, others (define the relationship):a) D-Ribose and L-Riboseb) D-Arabinose and D-ribulosec) D-mannose and D-talosed) 2-deoxy-α-D-ribose and D-ribosee) Glucose and maltosef) α-D-galactopyranose and α-D-galactofuranoseg) β-D-galactopyranose and α-D-galactopyranoseh) D-psicose and D-tagatoseProblem 41) Allolactose is a disaccharide similar to lactose that differ only in the position of the glyosidicbond (1,6 instead of 1,4). a) Explain whether allactose is a homo or heterodisaccharide?b) Draw the structure of lactose and allolactose and highlight the glycosidic bond. In bothstructures highlight the anomeric carbon (write it as mixture of the 2 anomers) and the nonreducingend. c) Allolactose can be a substrate of β-galactosidase as lactose. Draw the products of theenzymatic reactions (assuming mutarotation in aqueous solution). 2) Raffinose is a trisaccharide composed of α-D-galactose, D-glucose and D-fructose. It can befound in several vegetables including beans, cabbage, broccoli, asparagus, other vegetables, andwhole grains. Raffinose can be hydrolyzed to D-galactose and sucrose by the enzyme α- galactosidase, an enzyme not found in the human digestive tract.a) Draw the structure of raffinose and the product of the reactions of alpha-galactosidase (thehydrolyzed glycosidic bond is 1,6 - assume mutarotation in aqueous solution)b) After digesting of raffinose with alpha galactosidase the products of the reaction are treatedwith sucrase. Draw the final products of the reaction in the mixture in acqueous solution. c) Indicate which of the products are aldoses or ketoses.Problem 5Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) is a tetrameric enzyme that catalyzesone of the steps of glycolysis.A new colorimetric GAPDH assay provides a non-radioactive method to measure the enzymeactivity of GAPDH, based on the oxidation an alternative substrate, D-glyceraldehyde (note thatwith D-glyceraldehyde the enzyme activity is lower than the substrate D-glyceraldehyde3- phosphate). NADH (MW 633.4 g/mol) absorbs light at 340 nm while NAD+ does not show any absorption atthat wavelength.The Beer-Lambert Law relates the absorbance of a solute to its concentration:A = ε.c.LA is absorbance, ε is the molar extinction coefficient, c is concentration, and L is the path lengthof the solution in the spectrophotometer (usually in cm).In the lab you have available NADH. You dissolve 9.951 mg in 100 ml of buffer and measure anabsorbance of 0.4725 at 340 nm in a 0.5 cm cuvette.Subsequently in a 1 cm cuvette, you mix 25 µl of 20 mM NAD+, 25 µl of D-glyceraldehyde 0.2M , 400 µl of TAE (pH=8.6), and 50 µl of a solution containing GAPDH at a concentration of 12mg/l. You place the cuvette in a spectrophotometer and follow the kinetics of absorbance at 340nm. You measure a rate of decrease of absorbance (ΔA) of 0.43 min-1.a) Determine the absolute amount of NAD+ consumed after 30 s in the reaction (in nmol)b) Determine the specific activity of GAPDH in NADH produced per GADPH (inµmol/mg/min)Problem 6Certain Clostridium strains can anaerobically ferment pyruvate to ethanol, butanol and acetate(ABE fermentation) following this scheme:a) The fermentation pathway marked in red is called solvent fermentation phase. In thissolvent fermentation phase butanol, ethanol and acetone are generated. Explain the role of thesolvent fermentation phase as metabolic pathway in respect to glycolysis.b) What is the main difference in terms of metabolic energetics between solvent fermentationphase and the acid fermentation phase?Problem 7Ethanol is metabolized by alcohol dehydrogenase and aldehyde dehydrogenase to acetate, whichcan be susequently converted to acetyl-CoA.Ethylene glycol (an automotive antifreeze) is poisonous and oxidized to oxalic acid (as oxalate),a toxic compound found in rhubarb leaves and many other plants. Ethylene glycol is metabolizedby the same enzymes as ethanol, producing glycolate which is further oxidized to oxalate and notfurther metabolized.a) Write the reactions involved in Ethylene glycol conversion and explain also why itsmetabolism is not energetically favorable. b) Can ethylene glycol poisoning be prevented by intoxicating concentrations of ethanol? Ifyes, why?Problem 8Deficiency of pyruvate kinase in the red blood cells can result in insufficient absorption of theright amount of O2. Explain which intermediate of the glycolysis is responsible for the observedeffect and why? Draw the substrate and product of the enzymatic reaction generating thatintermediate.Problem 9A mutation in Phosphofructokinase causes the loss of the allosteric site for ATP. Which effect onglycolysis is expected? Explain in detail.Explain which of the following mutations would result in the opposite effect on the rate ofglycolysis in liver:a) a mutation causing the loss of the binding site for citrate in phosphofructokinaseb) a mutation causing the loss of the binding site for fructose 1,6-bisphosphate in pyruvatekinase.c) A mutation decreasing the activity of adenylate kinaseProblem 10In the laboratory you are performing an experiment by growing yeast (S. cerevisiae) in strictlyanaerobic conditions, using 14C labeled glucose as carbon source. You isolate the products of thefermentation after you give enough time for each intermediate in the pathway to become labeled.You are able to isolate ethanol exclusively labeled on C-1. a) Determine position
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