Biochemistry 461 Section I May 21 1998 Final Exam Prof Jason D Kahn Your Printed Name Your SS Your 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 to answer 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 be available only through MARS Possibly Useful Information Michaelis Menten equation v0 Vmax S KM S where Vmax kcat E t Type of inhibition Apparent KM Apparent Vmax Apparent Vmax KM Competitive a KM Vmax 1 a Vmax KM Uncompetitive 1 a KM 1 a Vmax Vmax KM Mixed a a KM 1 a Vmax 1 a Vmax KM Noncompetitive a a KM 1 a Vmax 1 a Vmax KM a 1 I KI a 1 I KI Henderson Hasselbach equation pH pKa log A HA DG DH TDS DG RTlnQ where Q has the form of an equilibrium constant RT 2500 J mole today 2 Biochemistry 461 Final 5 21 98 1 30 points Nucleic Acids Amino Acids Hydrogen Bonding Adenine and thymine are shown below H bonded as they are in a W C base pair H O CH3 N H A N dR N H N N H T H N N O dR a 10 pts On the structure above identify the major and minor groove sides of the base pair and label the hydrogen bond donors and acceptors in each groove b 10 pts Asparagine residues in DNA binding domains are often found to recognize adenine in duplex DNA Draw a plausible structure for the interaction of an Asn side chain with the major groove edge of adenine c 10 pts The Watson Crick arrangement is not the only way that A and T can base pair Draw an alternative arrangement of A and T that might be observed if the sugar phosphate backbone didn t hold the bases in place in a helix in other words a different way to hydrogen bond them There is space on the next page for your answer if you need it Biochemistry 461 Final 5 21 98 2 15 pts Hemoglobin and Allostery Hemoglobin binds CO2 although not at the heme iron The T state binds CO2 much better than the R state via a reversible covalent linkage a 7 pts What effect would increased levels of CO2 have on the p50 for O2 binding to Hb Explain your reasoning b 8 pts Where is CO2 high in the body and where is it low Thus what are two likely physiological roles for CO2 binding to Hb 3 4 Biochemistry 461 Final 5 21 98 3 35 points Serine Protease Mechanism pH The 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 first step is just the replacement of the alcohol leaving group with water There is no need to draw the Asp side chain His Asp H Ser O O O O H N N H H O R1 R2 H2O R2OH b 6 pts How does this example illustrate covalent catalysis Biochemistry 461 Final 5 21 98 As pH is varied the kcat for chymotrypsin is found to be directly proportional to the amount of the His57 in the catalytic triad which is in the His as opposed to HisH form while Kdiss for substrate is constant 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 Structure Many membrane proteins are attached in the hydrophobic interior of the lipid bilayer by one or more a helices which span the membrane a 8 pts How can we sometimes identify the trans membrane a helices by examination of the protein sequence even for a new protein with no homology to any others 5 Biochemistry 461 Final 5 21 98 b 8 pts When b sheets are found in membranes they are usually complete barrels not one or two strands Why contrast with a helices in membranes c 6 pts Soluble proteins which interact with membrane receptors are often anchored to the membrane 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 tailed detergents 6 7 Biochemistry 461 Final 5 21 98 CH2OH 5 30 points Carbohydrates Bioenergetics The Haworth projection for glucose is shown at the right a 6 pts Draw the Haworth projection for galactose the C 4 epimer of glucose O OH OH HO OH b 8 pts Draw the Haworth projection of lactose which is b DGalactopyranosyl 1 4 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 it much less important to specify the anomer of the glucose unit Biochemistry 461 Final 5 21 98 d 10 pts The enzyme b galactosidase hydrolyzes the b 1 4 linkage in lactose in the cell This enzyme has no cofactor requirements and does not use ATP just like proteases What does this simple observation tell us about the thermodynamic stability of the disaccharide relative to its constituent 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 Inhibition Irreversible affinity labels or mechanism based inhibitors react in an enzyme active site and thereby inactivate the enzyme a 6 pts Give one example of an irreversible inhibitor and the enzyme it inhibits if you don t know the names a description or structure will do b 5 pts Give one reason why irreversible enzyme inhibitors are important in contemporary society 8 Biochemistry 461 Final 5 21 98 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 protect an 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 inhibitor subsequent characterization of the Michaelis …