Fall 2017 Bio 317 Exam 2A Key

[All questions are 4 points each unless otherwise indicated]

1. You are trying to convert elastase into chymotrypsin by genetic engineering. A key
component of your strategy might be:
A. to replace the serine in the catalytic triad with tyrosine to strengthen specific binding of
aromatic substrates.
B. to enlarge the size of the oxyanion hole to accommodate aromatic substrates.
C. to introduce large aromatic amino acid side chains into the oxyanion hole to strengthen
specific binding of aromatic substrates.
D. to reduce the size of the side chains of the amino acids lining the specificity pocket.
E. to replace the histidine residue in the catalytic triad with glycine to increase the likelihood of a
nucleophilic attack on the substrate.

2. Which of the following statements about the catalytic mechanism of serine proteases is
TRUE?
A. The side chain of the catalytic triad aspartate residue engages in covalent catalysis by
carrying out a nucleophilic attack on the substrate.
B. The measured rate of peptide bond cleavage is likely to be the same at pH 2 and pH 10.
C. The measured rate of catalysis would be identical in an aqueous environment and a water-
free environment.
D. The ability of a serine residue to carry out a nucleophilic attack is strongly influenced
by the identity of a residue immediately adjacent in the tertiary structure.
E. The two final products of the serine protease reaction contain backbone carbonyl carbons
with tetrahedral geometries.

3. Chymotrypsin becomes activated from an inactive zymogen named chymotrypsinogen.

Based on your understanding of this activation, which of the following steps in the chymotrypsin
catalytic mechanism is most likely to be impaired before activation?
A. Formation of hydrogen bonds between peptide backbone atoms of the protease and a
charged atom of a reaction intermediate.
B. Entry of a water molecule into the active site.
C. Transformation of the catalytic triad serine residue into a nucleophile.
D. Polarization of the catalytic triad histidine residue by a neighboring aspartate residue.
E. Formation of an oxonium ion transition state intermediate.

4. You travel to Mars to compare Michaelis-Menten kinetics of enzymes originating from

Humans and Martians. Both enzymes catalyze the turnover of identical substrates and all
reactions were carried out with identical enzyme concentrations.

Enzyme Vmax (s-1) KM (nM) kcat/KM (s-1 nM-1)

Human 150 100 3
Martian 300 50 12

Which of the following conclusions based on these results are CORRECT?

A. The catalytic efficiency of the Human enzyme is greater than Martian enzyme.
B. The affinity of the Martian enzyme for the substrate is weaker than the Human enzyme.
C. The rate of the k2 step for the Martian enzyme is greater than the Human enzyme.
D. The overall reaction catalyzed by the Martian enzyme has a less negative ΔG value than the
reaction catalyzed by the Human enzyme.
E. The Human enzyme becomes saturated with substrate at lower substrate concentrations
than the Martian enzyme.
 Fall 2017 Bio 317 Exam 2A Key

5. Compare the two curves on the Michaelis-Menten plot below showing steady state kinetic
measurements of two different enzymes catalyzing the same reaction. Which of the following
statements is INCOMPATIBLE with the information shown on the plot?

A. The affinity of enzyme 1 for the substrate is greater than the affinity of enzyme 2 for the
substrate.
B. If the reactions used to generate both curves were carried out with the same concentration of
enzyme, the value of kcat for enzyme 2 would be lower than that for enzyme 1.
C. At saturating substrate concentrations, enzyme 1 catalyzes the conversion of substrate to
product at a higher rate.
D. If the same data were plotted on a Lineweaver-Burk plot, the fitted straight line for
enzyme 1 would intersect the y-axis at a higher value.
E. If the reactions used to generate both curves were carried out with the same concentration of
enzyme, the catalytic efficiency of enzyme 1 would be higher than the catalytic efficiency of
enzyme 2.

6. Which of the following is NOT a requirement for measuring Michelis-Menten kinetics?

A. The reaction must be at equilibrium.
B. The rate of the k2 step must be significantly greater than the rate of the k-1 step.
C. The reaction must be at steady-state.
D. The concentration of the enzyme substrate complex must not significantly change over time.
E. The concentration of substrate must be in excess over the concentration of enzyme.
 Fall 2017 Bio 317 Exam 2A Key

7. I propose to design a new drug that will act as an inhibitor for an enzyme. If I engineer it with
similar chemical properties to the transition state of the reaction, what type of inhibitor am I
attempting to engineer and how will I know if I have succeeded?
A. A competitive inhibitor, collect kinetic data both in the presence and absence of inhibitor and
watch for a decrease in Vmax.
B. A competitive inhibitor, collect kinetic data both in the presence and absence of
inhibitor and watch for an increase in KM.
C. A competitive inhibitor, collect kinetic data both in the presence and absence of inhibitor and
watch for a decrease in KM.
D. A mixed noncompetitive inhibitor, collect kinetic data both in the presence and absence of
inhibitor and watch for a decrease in KM.
E. A mixed noncompetitive inhibitor, collect kinetic data both in the presence and absence of
inhibitor and watch for an increase in Vmax.

8. Which of the following is the best explanation for the observed decrease in the Vmax of an
enzyme-catalyzed reaction in the presence of an uncompetitive inhibitor?
A. The inhibitor prevents binding of the substrate to the enzyme active site.
B. The inhibitor increases the rate of the k2 step.
C. The inhibitor alters the standard free energy of the substrate.
D. The inhibitor binds to the free enzyme (E) and deforms the active site to prevent catalysis.
E. Formation of the ESI complex depletes the amount of ES that is available to be
catalyzed into product.

9. Given below are kinetic parameters for an ATPase measured in the presence of increasing
concentrations of a small molecule that acts as an inhibitor to the enzyme. The enzyme
concentration was kept constant in all reactions.

Inhibitor kcat (min-1) KM for ATP (nM) kcat/KM (min-1 nM-1)

concentration (nM)
0 1250 130 9.6
20 800 130 6.2
50 600 130 4.6

According to the data on the ATPase given above, which is the most likely mechanism of
inhibition used by this small molecule?
A. Competitive inhibition through binding to the free enzyme at the substrate-binding site.
B. Uncompetitive inhibition through binding preferentially to the enzyme substrate complex.
C. Uncompetitive inhibition through binding preferentially to the free enzyme
D. Mixed noncompetitive inhibition through binding with high affinity to the free enzyme and with
low affinity to the enzyme-substrate complex.
E. Pure noncompetitive inhibition through binding with equal affinity to the free enzyme
and enzyme-substrate complex.

10. If defective myosin head groups were able to bind and hydrolyze ATP but not able to
release inorganic phosphate (Pi) after hydrolysis, which step in the actin-myosin cycle would
NOT take place?
A. detachment of the myosin head group from the actin filament.
B. cocking forward of the myosin head group while detached from the actin filament.
C. weak attachment of the myosin head group to the actin filament.
D. strong attachment of the myosin head groups to the actin filament.
E. the conformational change in the myosin head group in the presence of Ca2+.
 Fall 2017 Bio 317 Exam 2A Key

11. Calcium ions are released into muscle fibers from the sarcoplasmic reticulum to regulate
contraction. What would be the most likely consequence of a defective sarcoplasmic reticulum
that could not release calcium?
A. The thick filaments of the sarcomeres would be unable to form cross-bridges with the
thin filaments.
B. Hydrolysis of ATP by the myosin head would occur at a faster rate.
C. The F-actin filaments would disassemble into G-actin monomers.
D. The position of the tropomyosin ‘cable’ on the thin filament would not change in response to
ATP hydrolysis by troponin.
E. The transient intermediate formed immediately before the “power stroke” would persist for a
longer time.

12. The ATP hydrolysis activity of actin is used to:

A. assemble actin filaments.
B. disassemble actin filaments.
C. drive actin-myosin interactions.
D. break actin-myosin interactions.
E. contract sarcomeres.

13. You are working as a scientist in an antibody-design company. Which alterations to a

monomeric antibody would have the greatest effect on its antigen-binding specificity?
A. Swap the C-terminal ~110 amino acids of the heavy chains with those from a different
antibody.
B. Swap the C-terminal immunoglobulin fold domain of the light chains with those from a
different antibody.
C. Alter the primary structures of the loops connecting the beta strands at the N-terminal
ends of both the heavy and light chains.
D. Add an additional immunoglobulin fold domain at the C-terminus of the heavy chain.
E. Swap the N-terminal immununglobulin fold domains of the two light chains within the same
antibody.

14. IgM is one of five classes of antibodies that can be produced by our immune system. Which
of the following is a possible subunit composition of IgM?
A. 10 μ-chains, 10 λ-chains and a J chain.
B. 5 κ-chains, 5 λ-chains, and a J chain.
C. 2 α-chains, and 2 λ-chains.
D. 4 μ-chains, 4 κ-chains, a J chain and a secretory component.
E. 5 δ-chains and 5 κ-chains.

15. Which of the following statements about antibody binding is TRUE?

A. The heavy and light chains of an antibody bind together using only noncovalent interactions.
B. Antibodies that have been in unfolded in high concentrations of urea will bind to their target
antigen with identical affinity as the folded antibody.
C. Replacing the heavy chains of an antibody with the heavy chains of a different
antibody from the same class would affect the specificity of antigen binding.
D. Antibodies contain two heavy chains made up of constant domains and two light chains
made up of variable domains.
E. Cleavage of IgG with pepsin generates two Fab fragments and an Fc fragment.
 Fall 2017 Bio 317 Exam 2A Key

16. You have created a library of variant hemoglobins. One variant contains hemes with a
previously unknown metal that replaces the iron. The hemes have been reported to bind oxygen
reversibly but the diameter of the metal is too small to alter the position of the F helix when
oxygen is bound or released. What is the most likely consequence of this metal substitution?
A. The hemoglobin might display an S-shaped oxygen binding curve with markedly increased
affinity
B. The hemoglobin might display an S-shaped oxygen binding curve with markedly decreased
affinity.
C. The hemoglobin might display a “normal” S-shaped oxygen binding curve like that of human
hemoglobin A, but it would have no Bohr effect.
D. The hemoglobin would have a strong tendency to dissociate into monomeric subunits if the F
helix no longer shifts.
E. If the F helix does not move, the oxygen binding curve would most likely be hyperbolic
with no Bohr effect.

17. Another hemoglobin variant has the C-terminal residues of both alpha and beta chains
replaced by glycines. What is the most likely consequence of these amino acid substitutions?
A. The hemoglobin would have a strong tendency to dissociate into monomeric subunits if the
C-termini were replaced by glycines.
B. Since glycine is a small amino acid, the substitution might not affect either the shape of the
oxygen binding curve or the Bohr effect.
C. The substitution might alter but not eliminate the S-shape of the oxygen dissociation
curve but it would most likely eliminate the Bohr effect.
D. The substitution by such a small amino acid would result in slippage of the hemes out of their
pockets and loss of reversible oxygen binding.
E. The small amino acid might facilitate spontaneous polymerization of the hemoglobin even
without any other amino acid substitutions.

18. Carbon dioxide can react with the amino termini of hemoglobin A to form
carbaminohemoglobin, but cannot react with the amino termini of hemoglobin F, which are
acetylated. What is the consequence of loss of CO2 binding to the blocked amino termini of the
alpha chains of hemoglobin F?
A. Since only a relatively small fraction of CO2 is carried by the amino termini, the effect would
be modest,and all of the protons derived from reaction of CO2 with water could still be carried
as Bohr protons by the beta or gamma chains.
B. The contribution of CO2 to the blood is not nearly as great as the contribution of bicarbonate
ions generated by metabolism, so the major effect on hemoglobin would be that of failure of
bicarbonate ions, not carbon dioxide, to react directly with the amino termini of the protein.
C. The amino termini of the alpha chains participate in ion pairs, so if they are blocked with CO2
the protein’s total capacity to bind oxygen would be reduced.
D. The amino termini of the alpha chains participate in oxygenation-linked ion pairs, so if
they are blocked in hemoglobin F, there would be not ony a further decrease in modest
binding of CO2 but also an increase in affinity for oxygen.
E. The amino termini of the alpha chains participate in oxygenation-linked ion pairs, so if they
are blocked in hemoglobin F, there would be not ony a further decrease in modest binding of
CO2 but also a decrease in affinity for oxygen.
 Fall 2017 Bio 317 Exam 2A Key

19-22. (2 points each) For the following observations answer A if the observation is consistent
only with the application of the Monod, Wyman, Changeux model of cooperativity to the stated
properties of hemoglobin and myoglobin, answer B if the observation is consistent only with the
application of the Koshland, Nemethy Filmer model to the stated properties of hemoglobin and
myoglobin, answer C if both models apply to the stated properties of hemoglobin and
myoglobin, and answer D if neither model is applicable to the stated properties of hemoglobin
and myoglobin. (The properties of hemoglobin and myoglobin as stated are all correct.)

19 __C____ Fully oxygenated hemoglobin has a different conformation than fully deoxygenated
hemoglobin.
20.___D___ Myoglobin binds oxygen reversibly.
21.___B___ As hemoglobin binds each of four molecules of oxygen, it appears that as each
oxygen is bound a proton is simultaneously released.
22.__A____ Attempts to detect multiple conformational species of hemoglobin in solution
inevitably reveal only mixtures of two conformational species.

23. You are studying the synthesis of a protein that is O-glycosylated. Which statement is
INCORRECT
A. A so-called “consensus sequence” in the polypeptide chain determines the
glycosylation sites.
B. The complete oligosaccharide chain is synthesized in the cytosol and then transferred to the
polypeptide chain in the Golgi for glycosylation.
C. The sugars are attached only after synthesis of the polypeptide chain is completed.
D. Different enzymes are required to put on different sugars.
E. The glycosylation events are considered to be “post-translational” rather than “co-
translational, and take place on serines or threonines.

24. Which statement about the effects of penicillin and beta-lactamases on bacterial cell walls is
correct?
A. penicillin blocks formation of pentaglycine bridges and beta-lactamases block formation of
tetrapeptide bridges in the bacterial cell wall.
B. penicillin blocks formation of tetrapeptide bridges and beta-lactamases block formation of
pentaglycine bridges in the bacterial cell wall.
C. penicillin blocks cleavage of the NAG-NAM co-polymeric polysaccharide structure of the cell
wall and beta lactamases catalyze the cleavage of the tetrapeptide bridges in the wall.
D. penicillin blocks formation of the pentaglycine bridges and beta lactamases cleave the
four-membered ring in penicillin.
E. None of these statements would fully account for the effects of both penicillin and beta-
lactamases on antimicrobial activity.
 Fall 2017 Bio 317 Exam 2A Key

25. In glycogen, both 1-4 and 1-6 linkages between glucose residues can be identified. Which
statement about these linkages is true?
A. Both linkages would eliminate the possilibility of any reaction of the products of these
linkages with a reagent that can detect a “reducing end” in the products.
B. The product of the 1-4 linkage in glycogen would still react with a reagent that can detect a
“reducing end” in the product, but the product of the 1-6 linkage could not react with the reagent.
C. The product of the 1-6 linkage in glycogen would still react with a reagent that can detect a
“reducing end” in the product, but the product of the 1-4 linkage could not react with the reagent.
D. Both linkages would diminish but not eliminate the yield of reaction with a reagent that
can detect a “reducing end” in the products.
E. Linkage of glucose molecules through 1-4 or 1-6 linkages as found in glycogen would have
no effect on the number of “reducing ends” that could be detected with a reagent that can detect
such reducing ends.

26. Which statement about regulation of metabolic pathways is most correct?

A. The most distinctive feature of a step in a metabolic pathway that makes it likely to be “rate
limiting” is that it is the first step in the pathway.
B. The most distinctive feature of a step in a metabolic pathway that makes it likely to be “rate
limiting” is that it is the last step in the pathway.
C. The most distinctive feature of a step in a metabolic pathway that makes it likely to be
“rate limiting” is that the concentrations of reactants and products are not at their
equilibrium values.
D. The most distinctive feature of a step in a metabolic pathway that makes it likely to be “rate
limiting” is that the enzyme catalyzing the reaction is almost always tetrameric.
E. The most distinctive feature of a step in a metabolic pathway that makes it likely to be “rate
limiting” is that the enzyme catalyzing the reaction conforms to the MWC model of allostery.

27. You are examining the characteristics of a pathway in which the first step involves reaction
of ATP with an alcohol to form a phospho-ester and the inorganic phosphoanhydride called
sodium pyrophosphate (the molecule contains two phosphates linked through a
phosphoanhydride linkage); the second step involves hydrolysis of sodium pyrophosphate to
form two sodium monophosphate molecules. Which description is most likely applicable to this
little pathway.
A. The first step is extremely favorable but the second step is extremely unfavorable.
B. The first step is probably somewhat favorable but the second step is extremely favorable.
C. The products of the pathway are most likely to be found in equilibrium with the reactants, with
no preferential direction for the pathway.
D. Neither of the reactions is encountered in biological systems because the chemistry of
phosphoryl compounds is inconsistent with the sequence described.
E. The pathway as described is most likely to describe how ATP is synthesized from inorganic
pyrophosphate.
 Fall 2017 Bio 317 Exam 2A Key

28.-32. (2 points each) The Monod-Wyman-Changeux and Koshland-Nemethy-Filmer models

of cooperativity have been used extensively to explain the properties of many enzymes that are
highly regulated [think phosphofructokinase here] in addition to hemoglobin. Indicate whether
the observation is most consistent with the MWC model, the KNF model, both models, or
neither model by using the following key: A. only MWC, B. only KNF, C. both MWC and KNF, D.
neither MWC nor KNF.
28. ___B____ Enzyme I is a tetramer with four substrate binding sites that contains four small
molecules in the absence of substrate. Each time a molecule of substrate binds, a small
molecule is liberated, but no species such as a form in which three small molecules have been
liberated but only one substrate molecule is bound [this scenario would be an example of the
observation Rbar does not equal Ybar, which applies only to the MWC model] can ever be
found.
29. ___D____ Enzyme II is also a tetramer with four substrate binding sites and a markedly
sigmoidal substrate binding curve. Graphical analysis of substrate binding to Enzyme II using a
Hill plot gives a value of nH of 6.5 [nH can’t exceed the number of subunits.].
30. ___C____ Enzyme III is also a tetramer with four substrate binding sites that can be
dissociated into dimers. Graphical analysis of substrate binding to Enzyme III using a Hill plot
gives a value of nH of 3.0 under conditions when the enzyme remains tetrameric but the value
of nH falls to 1.5 when the enzyme is split into dimers [both models can explain positive
cooperativity in tetramers and dimers].
31. ___C____ The affinity of Enzyme IV for substrate can be regulated by analogs of the small
molecule that bind with greater affinity to the enzyme when substrate is bound than when the
substrate is absent [both models are consistent with the effects of a positive allosteric effector,
such as AMP binding to PFK].
32. ___C____ The affinity of Enzyme V for substrate can regulated by analogs of the small
molecule that bind with greater affinity to the enzyme when the substrate is absent than when
the substrate is bound [both models are also consistent with the effects of a negative allosteric
effector, such as ATP binding to PFK].