Name:_________________________________

SAMPLE Test 2C430, BiochemistryApril 10, 2013

Problem 1 [3 points] SDS-PAGE separates proteins primarily due to differences in:

A) isoelectronic pointsB) sizeC) polarityD) solubilityE) sequence

Answer:

Problem 2[3 points] Which of the following proteins would be last to elute at high pH from an anion-exchange column?

A) Protein A (pI = 5)B) Protein B (pI = 7)C) Protein C (pI = 9)D) All are retainedE) None are retained

Answer:

Problem 3[3 points] Which of the following statements does not apply to the K value in the equation for the oxygen binding curve of myoglobin?

A) It is numerically equal to p50.B) It is defined as that oxygen partial pressure at which half of the oxygen binding sites are occupied.C) It is a measure of the affinity of myoglobin for oxygen.D) If Y > K, then myoglobin is less than 50% saturated with oxygen.E) It is the value of pO2 at which Y = 0.5.

Answer: D?

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Problem 4[3 points] Consider a hypothetical hemoglobin with a Hill coefficient of 1 and the same p50 value as normal hemoglobin. Choose the statement below that best describes the two proteins.

A) There is a cooperative interaction between oxygen-binding sites in both the hypothetical and normal hemoglobins.B) The hypothetical hemoglobin has a greater oxygen affinity than normal hemoglobin.C) The oxygen binding curve for the hypothetical hemoglobin is hyperbolic, and the curve for normal hemoglobin is sigmoidal.D) The two hemoglobins would be able to deliver about the same amount of oxygen to the tissues.E) At pO2 less than p50, normal hemoglobin has a greater YO2 value.

Answer: C?

Problem 5[3 points] Which of the following increases the affinity of hemoglobin for oxygen:

A) an increase in BPG concentration.B) the formation of N-terminal carbamates.C) increasing pH.D) all of the aboveE) none of the above

Answer: C

Problem 6[3 points] Which of the following is true for the symmetry model of allosterism:

A) the protein is an oligomer of symmetrically (or pseudosymmetrically) related subunits.B) the oligomer can exist in two conformational states which are in equilibrium.C) the ligand can bind to a subunit in either conformation.D) the molecular symmetry of the protein is conserved during the conformational change.E) all of the above.

Answer: E

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Problem 7[3 points] During the T to R conformational shift, Fe(II) drags the F-helix via a bond to the side chain of ________.

A) Leu F7B) Leu F4C) His F8D) Leu FG3E) Val FG5

Answer: C

Problem 8[3 points] An enzyme is considered to have evolved to its most efficient form if

A) kcat is a large numberB) kcat/KM is near the diffusion-controlled limitC) KM is a large numberD) kcat/KM is a very small numberE) KM is a small number

Answer: B

Problem 9[3 points] When [S] = KM, 0 = _____Vmax.

A) 2B) 1C) 0.67D) 0.5E) 0.1

Answer: D?

Problem 10[3 points] Parallel lines on a Lineweaver-Burk plot are diagnostic of:

A) competitive inhibition.B) non-competitive inhibition.C) allosteric activation.D) allosteric inhibition.E) uncompetitive inhibition.

Answer: E

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Problems 11 and 12 refer to the overall transformation:

Problem 11[3 points] For the reaction, the steady state assumption assumes thatA) [S] = [P]B) [ES] is constantC) [P]>>[E]D) [P] is constantE) k–1>>k2

Answer: B?

Problem 12[3 points] The Michaelis constant KM is defined asA) (k–1 + k2)/k1

B) (k–1 + k1)/k2

C) ([P] + [E])/[ES]D) [ES]E) none of the above

Answer: A

Problem 13[3 points] A compound that reduces the concentration of enzyme available for substrate binding is called:

A) a transition-state analogB) a non-competitive inhibitorC) an allosteric effectorD) an enzyme inactivatorE) a competitive inhibitor

Answer: E

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Problems 14 and 15 refer to the diagram (with boxes where it has been left incomplete):

Problem 14[3 points] This diagram refers to a:

A) Ping Pong reaction.B) ordered bisubstrate reaction.C) random bisubstrate reaction.D) double displacement reaction.E) not enough information is provided.

Answer: C

Problem 15[3 points] Which reactant/product should be in the box labeled with a Z?

A) AB) BC) PD) QE) E

Answer: D

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Problem 16[3 points] Which of the following is not true about enzymes?

A) Enzymes catalyze reactions in the forward direction only.B) Enzymes can be very specific for their substrates.C) Enzyme activities can often be regulated.D) Enzymes typically act under milder conditions of temperature and pH than chemical catalysts.E) Enzymes typically catalyze reactions at much higher rates and at milder conditions than chemical catalysts.

Answer: A

Problem 17

[3 points] A pH/rate curve with an inflection point at pH~4 suggests the involvement of a(n) ________ in the catalytic step.

A) hydrophobic amino acidB) acidic amino acidC) basic amino acidD) metal ionE) redox cofactor

Answer: B

Problem 18[6 points] In order for an enzymatic reaction obeying the Michaelis-Menten equation to reach 3/4 of its maximum velocity, [S] would need to be how many times the Km? Show your work.

Answer:Km=[S]{Vm/V1 -1} The MnM equation is V = {V(max) *[S]} / {Km + [S]}

Now we have to determine [S] when V = 3/4 V(max). So lets put that in: 3/4 V(max) = {V(max) *[S]} / {Km + [S]}

For clarity, lets write the LHS as: 3 V(max) / 4 = {V(max) *[S]} / {Km + [S]}

Now lets invert both sides of the equation: 4 / 3 V(max) = {Km + [S]} / {V(max) *[S]} = (Km / {V(max) *[S]}) + ([S] / {V(max) *[S]}) ... (Splitting the numerator) = (Km / {V(max) *[S]}) + 1 / V(max) ... (Cancelling out [S])

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So, [4 / 3 V(max)] - [1 / V(max)] = (Km / {V(max) *[S]}) ... (Subtracting 1 / V(max) from both sides)

So, [4 / 3 V(max)] - [3 / 3V(max)] = (Km / {V(max) *[S]})

So, 1 / 3V(max) = (Km / {V(max) *[S]})

So, inverting the equation again: 3 V(max) = ({V(max) *[S]} / Km)

Divide both sides by V(max): 3 = [S] / Km

So, multiplying both sides by Km: 3 Km = [S]hence if V1 is 3/4 of Vm then for an enzymatic reaction obeying it to reach this velocity the [S] will be 3Km Hence d) option will be correct

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Problem 19[4 points] Consider the following hypothetical saturation data:

Log pO2 Log YO2/(1- YO2)12345

02.55

7.510

What is the Hill constant of this system? What is the effect of O2 binding to subsequent O2 binding? You do not need to plot the information for credit.

Answer:

Hill constant = slope = 5/2 = 2.5 Positive cooperativity because the hill constant >1

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Problem 20[6 points] The kinetics of E. coli ATCase were examined using increasing concentrations of aspartate, in the presence and absence of CTP and ATP, as shown below:

Looking at the shapes of the curves in this figure, does ATCase display typical Michaelis-Menten behavior in the presence of CTP? What kinetic parameter(s) appear to be changing in the presence of ATP? Is the reaction faster or slower at 5 mM aspartate in the presence of CTP?

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Problem 21 [23 points] Review the following except from the paper of Michihara et al. on the Purification of Mouse Mevalonate Pyrophosphate Decarboxylase (abbreviated MPD, Biol. Pharm. Bull. (2002), 25, 302-306).

a) What is the cellular location of mevalonate pyrophosphate decarboxylase in mouse liver?

b) The use of an MPD antibody-Affigel column indicates what type of chromatography?

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c) Why is the absorbance of the wash eluent checked at 280 nm?

d) How is mevalonate pyrophosphate decarboxylate eluted from the column?

e) Based on the graphs presented, what is the subunit composition of native MPD?

f) Provide values for A-D in the purification table of MPD, shown below:

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