WRITE YOUR NAME AND I.D. NUMBER LEGIBLY ON EVERY PAGE – PAGES WILL BE SEPARATED FOR GRADING! CHECK TO BE SURE YOU HAVE 6 PAGES, NAME (print): INCLUDING COVER PAGE. 5-digit course ID # I swear/affirm that I have neither given nor received any assistance with this exam. Signature: Date: A NON-PROGRAMMABLE CALCULATOR MAY BE USED ON THIS EXAM. No programmable calculators are permitted, and no sharing of calculators. We have a couple of spare calculators to lend in an emergency. SHOW YOUR WORK FOR ALL CALCULATIONS, AND BE SURE TO STATE UNITS OF ANY NUMERICAL ANSWERS. If the reasoning, calculations, or answer are shown anywhere other than in the space provided, make a note in the space provided and put answer on BACK OF SAME PAGE so the grader for that page will have it. USEFUL CONSTANTS: R (gas constant) = 8.315 J•mol–1•Kelvin–1 = 8.315 x 10–3 kJ•mol–1•Kelvin–1 If temperature = 25 °C, absolute temperature T = 298 K (Assume this temperature unless problem states otherwise.) BIOCHEMISTRY 460 FIRST HOUR EXAMINATION FORM A (yellow) ANSWER KEY February 6, 2006 Use these “generic” pKa values only when no precise pKa for a specific group is given. Ionizable group in peptides and proteins Approximate ("generic") pKa in peptides & proteins (from Berg, Tymoczko & Stryer, Biochemistry, 5th ed., 2001) α-carboxyl 3.1 side chain carboxyl 4.1 imidazole 6.0 α-amino 8.0 thiol 8.3 aromatic hydroxyl 10.9 ε-amino 10.8 guanidino 12.5Biochemistry 460, Exam #1 Form A ANSWERS NAME February 6, 2006 5-digit course ID# page 2 1. (31 pts) A tetrapeptide structure is shown below: A. (4 pts) Draw one clear ARROW ( ↑ ) to any ONE of the peptide bonds in this structure and label it “peptide bond”. Label only one peptide bond. (Any one blue arrow on structure) B. (4 pts) Circle ONE group of 6 atoms in the tetrapeptide that are coplanar (all in the same plane), excluding atoms in cyclic structures. There is more than one correct answer, but circle only 6 atoms total. (Any one red parallelogram on structure, or enclosed atoms could be circled separately) C. (4 pts) Draw a labeled arrow to ONE bond whose rotation angle is a φ angle (label that arrow φ) and another arrow to ONE bond whose rotation angle is a ψ angle (label that arrow ψ). Φ: labeled arrow can be drawn to any amide N–Cα bond; ψ: arrow to any Cα–carbonyl C bond (no credit for arrows to atoms; arrows must be to BONDS.) D. (6 pts) Identify the residues in the peptide above, using 3-letter abbreviations: (1) (2) (3) (4) _Arg – _Tyr – _His – _Val E. (4 pts) At which of the following pH values would this peptide exist predominantly in the state of ionization shown above? (Assume "generic" pKas of functional groups in peptides and proteins shown on cover sheet of this exam.) a) pH 1 b) pH 3.5 c) pH 5 *d) pH 7.5 e) pH 10 f) pH 13 F. (6 pts) At pH 11.5, what would be the charges (if any) on the terminal groups and the side chain (R group) of each residue in this peptide: positively charged (+), negatively charged (–), or uncharged (0)? What would be the net charge on this peptide at pH 11.5? !functional group Charge, if any, at pH 11.5 ( – , + , or 0 ) N-terminal amino group of chain 0 C-terminal carboxyl group of chain – R group (side chain) of residue 1 + R group (side chain) of residue 2 – R group (side chain) of residue 3 0 R group (side chain) of residue 4 0 NET charge at pH 11.5: –1 G. (3 pts) At pH 7, would the side chain of residue 2 on the peptide above be able to act as a hydrogen bond donor or as a hydrogen bond acceptor or both? ______BOTH________ /31 p. 2 (31 points) p. 3 (20 points) p. 4 (17 points) p. 5 (20 points) p. 6 (12 points) TOTAL: (100 points)Biochemistry 460, Exam #1 Form A ANSWERS NAME February 6, 2006 5-digit course ID# page 3 2. (12 pts) The catalytic activity of the enzyme lysozyme requires that the side chain of a specific glutamate residue be in its neutral form for the enzyme to be active. That specific Glu side chain in the protein has an unusually high pKa, about 5.9. To receive credit, you must show your work for all calculations. A. (5 pts) For a functional group with a pKa of 5.9, what is the base/acid ratio at pH 5.4? pH = pKa + log [base]/[acid] log [base]/[acid] = pH – pKa = 5.4 – 5.9 = –0.5 [base]/[acid] ratio = 10–0.5 = 0.316 / 1 B. (3 pts) Is it the conjugate base or the conjugate acid form of the Glu side chain that is the catalytically active form? Catalytically active form is conjugate ACID (neutral, R-COOH) C. (4 pts) What fraction of the total enzyme molecules in a solution of lysozyme at pH 5.4 have that Glu side chain in the active form? Active/total = [acid] / ([acid] + [base]) = 1 / (1 + 0.316) = 0.76 or 76% active (neutral) 3. (8 pts) Suppose you have a mixture of 5 proteins (A-E) with the properties described below. Protein isoelectric point (pI) subunit M.W. native M.W. A 5 20,000 80,000 B 3 50,000 200,000 C 7 75,000 75,000 D 9 30,000 120,000 E 8 15,000 and 45,000 (2 stained bands) 60,000 A. (4 pts) If you were separating a mixture of these proteins on a cation exchange column (– charged groups on column material, e.g., carboxymethyl– groups) at pH 6.0, which protein would elute (come off the column) LAST? __D__ (protein with greatest net positive charge at pH 6, which is protein with highest pI) B. (4 pts) If you were separating a mixture of these proteins on a molecular sieve column under physiological (non-denaturing) conditions, which protein would elute FIRST? __B__ (protein with the highest native M.W.) /20Biochemistry 460, Exam #1 Form A ANSWERS NAME February 6, 2006 5-digit course ID# page 4 4. (17 pts) In a muscle cell, when the ATP required to provide the energy for muscle contraction is depleted, another "energy storage" compound, creatine phosphate, can provide the energy to produce replenish the supply of ATP from ADP in the following reaction: ADP + creatine phosphate ↔ ATP + creatine A. (4 pts) Write the expression defining the equilibrium constant for this reaction in terms of the concentrations of the reactants and products at equilibrium. What would be the
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