Biochemistry 461, Section I Your Printed Name: May 21, 1998Final Exam Your SS#: Prof. Jason D. KahnYour Signature: You have 120 minutes for this exam.The exam has 7 questions, worth 200 points. Do all 7 questions. In many cases, you do not need toanswer earlier parts of questions to answer the later ones.Exams written in pencil or erasable ink will not be re-graded under any circumstances.Explanations should be concise and answer the specific question asked.You will need a calculator for this exam. No other study aids or materials are permitted.There will be a viewing at a time and place to be announced on the class web page. Final grades will beavailable only through MARS.Possibly Useful Information:Michaelis-Menten equation: v0 = Vmax[S]/(KM + [S]), where Vmax = kcat[E]tType of inhibitionApparent KMApparent VmaxApparent Vmax /KMCompetitivea KMVmax(1/a) Vmax/KMUncompetitive(1/a¢) KM(1/a¢) VmaxVmax/KMMixed(a/a¢)KM(1/a¢) Vmax(1/a) Vmax/KMNoncompetitive (a=a¢)KM(1/a) Vmax(1/a) Vmax/KMa = 1 + [I]/ KIa¢ = 1 + [I]/ KI¢Henderson-Hasselbach equation: pH = pKa + log([A–]/[HA])DG = DH - TDS = DG°¢ + RTlnQ, where Q has the form of an equilibrium constantRT = 2500 J/mole todayBiochemistry 461 Final 5/21/9821. (30 points) Nucleic Acids, Amino Acids, Hydrogen BondingAdenine and thymine are shown below, H-bonded as they are in a W-C base pair.(a; 10 pts) On the structure above, identify the major and minor groove sides of the base pair, andlabel the hydrogen bond donors and acceptors in each groove.(b; 10 pts) Asparagine residues in DNA binding domains are often found to recognize adenine induplex DNA. Draw a plausible structure for the interaction of an Asn side chain with the majorgroove edge of adenine.(c;10 pts) The Watson-Crick arrangement is not the only way that A and T can base pair. Draw analternative arrangement of A and T that might be observed if the sugar-phosphate backbone didn’thold the bases in place in a helix (in other words, a different way to hydrogen-bond them). There isspace on the next page for your answer if you need it.NNNNNNNOOdRdRCH3HHHHHATBiochemistry 461 Final 5/21/9832. (15 pts) Hemoglobin and Allostery.Hemoglobin binds CO2, although not at the heme iron. The T state binds CO2 much better than the Rstate (via a reversible covalent linkage).(a; 7 pts) What effect would increased levels of CO2 have on the p50 for O2 binding to Hb? Explainyour reasoning.(b; 8 pts) Where is CO2 high in the body and where is it low? Thus, what are two likely physiologicalroles for CO2 binding to Hb?Biochemistry 461 Final 5/21/9843. (35 points) Serine Protease Mechanism, pHThe structure below is the acyl-enzyme intermediate in the hydrolysis of an ester by a serine protease,which has a mechanism similar to the amide hydrolysis we have studied.(a; 16 pts) Draw the structures for the next two steps, paying attention to protonation states. The firststep is just the replacement of the alcohol leaving group with water. There is no need to draw the Aspside chain.(b; 6 pts) How does this example illustrate covalent catalysis?SerOOR1OR2HN NHisAspOOHHHH2OR2OHBiochemistry 461 Final 5/21/985As pH is varied, the kcat for chymotrypsin is found to be directly proportional to the amount of theHis57 in the catalytic triad which is in the His as opposed to HisH+ form, while Kdiss for substrate isconstant. The pKa for His57 is 7.2.(c; 5 pts) Is the histidine acting as a general acid or as a general base? How do you know?(d; 8 pts) Calculate the ratio of kcat at pH 6.7 to kcat at pH 7.7; in other words, calculate[His]/([His] + [HisH+]) at the two pH’s and take the ratio of the answers.4. (30 pts) Biomembranes, Protein StructureMany membrane proteins are attached in the hydrophobic interior of the lipid bilayer by one or morea helices which span the membrane.(a; 8 pts) How can we sometimes identify the trans-membrane a helices by examination of theprotein sequence, even for a new protein with no homology to any others?Biochemistry 461 Final 5/21/986(b; 8 pts) When b sheets are found in membranes, they are usually complete barrels, not one or twostrands. Why (contrast with a helices in membranes)?(c; 6 pts) Soluble proteins which interact with membrane receptors are often anchored to themembrane by hydrophobic lipid tails. How does this improve the efficiency of signal transduction?(d; 8pts) Why are biomembranes made of two-tailed phospholipids rather than single-taileddetergents?Biochemistry 461 Final 5/21/9875. (30 points) Carbohydrates, BioenergeticsThe Haworth projection for glucose is shown at theright.(a; 6 pts) Draw the Haworth projection for galactose,the C-4 epimer of glucose.(b; 8 pts) Draw the Haworth projection of lactose, which is b-D-Galactopyranosyl(1Æ4)Glucopyranose. Circle the two anomeric carbons.(c; 6 pts) Why are we careful to specify which anomer of galactose is present on lactose, but find itmuch less important to specify the anomer of the glucose unit?OHOOHOHCH2OHOHBiochemistry 461 Final 5/21/988(d; 10 pts) The enzyme b-galactosidase hydrolyzes the b(1Æ4) linkage in lactose in the cell. Thisenzyme has no cofactor requirements and does not use ATP, just like proteases. What does thissimple observation tell us about the thermodynamic stability of the disaccharide relative to itsconstituent monosaccharides? Thus, without being able to say anything about the detailed pathway,what must the cell do in order to synthesize polysaccharides?6. (35 points) Michaelis-Menten Kinetics and InhibitionIrreversible affinity labels or mechanism based inhibitors react in an enzyme active site and therebyinactivate the enzyme.(a; 6 pts) Give one example of an irreversible inhibitor and the enzyme it inhibits (if you don’t knowthe names, a description or structure will do).(b; 5 pts) Give one reason why irreversible enzyme inhibitors are important in contemporary society.Biochemistry 461 Final 5/21/989(c; 8 pts) Why do irreversible inhibitors usually resemble substrates or transition states?(d; 8 pts) Based on your answer to (c), explain why a high concentration of true substrate can protectan enzyme from inactivation by an irreversible inhibitor. What kind of inhibition kinetics (non-, un-,or just plain competitive) does this remind you of?(e; 8 pts) In fact, if an enzyme is partially inactivated by treatment with an irreversible inhibitor(without substrate) for a short period of time followed by removal of the