MCB 354 Exam l

Some mutant viruses have arisen with decreased sensitivity to zanamivir. One such mutation is R118K within the neuraminidase active site. Draw the amino acids arginine and lysine at pH 7. What is the effect of this mutation?

Lysine is shorter than arginine and will no longer be sufficiently close to the carboxylate of zanamivir to hydrogen bond.

The neighboring residue at position 116 is also lysine. Explain how this perturbs the pKa of the catalytic lysine residue.

Two adjacent positive charges experience electrostatic repulsion. This destabilizes the ionization state in which both lysines are protonated, shifting the equilibrium for at least one lysine towards the neutral form (i.e. reduces the pKa and makes it a stronger acid).

Lys-116 is mutated to glutamate. How is the pKa of catalytic Lys-115 expected to change and why?

Negatively charged glutamate will have an attractive electrostatic interaction with protonated Lys-115, stabilizing the protonated species. The pKa for Lys-115 will therefore increase tremendously and it will become a stronger base.

In x-ray studies of crystalline peptides, Linus Pauling and Robert Corey found that the C−NC−N bond in the peptide link is intermediate in length (1.32 Å1.32 Å) between a typical C−NC−N single bond (1.49 Å1.49 Å) and a C=N double bond (1.27 Å1.27 Å). They also found that the peptide bond is planar (all four atoms attached to the C−NC−N group are located in the same plane) and that the two 𝛼-carbon-carbon atoms attached to the C−NC−N are always trans to each other (on opposite sides of the peptide bond).

a. What does the length of the C−NC−N bond in the peptide linkage indicate about its strength (i.e., whether it is single, double, or triple)?

b. What do the observations of Pauling and Corey tell us about the ease of rotation about the C−NC−N peptide bond?

(a) The C−N peptide bond can participate in resonance, or electron movement, between nitrogen and carbon, as well as the oxygen attached to the carbon in the C−N bond. Therefore, the C−N peptide bond has partial double-bond character, being shorter and stronger than a C−N single bond. However, the C−N peptide bond is longer and weaker than a C=N double bond.

(b) Since the C−N peptide bond undergoes resonance and has partial double-bond character, the bond cannot rotate like a single bond. Therefore, atoms associated with the C−N peptide bond are consistently found in a planar arrangement. The peptide bond does not contribute to either the 𝜙 or 𝜓 angles. The 𝜙 angle is associated with the single bond between nitrogen in the C−N peptide bond and the 𝛼 carbon of the peptide backbone. The 𝜓 angle is associated with the single bond between the carbon in the C−N peptide bond and the 𝛼 carbon of the peptide backbone.

William Astbury discovered that the x-ray diffraction pattern of wool shows a repeating structural unit spaced about 5.2 Å along the length of the wool fiber. When he steamed and stretched the wool, the x-ray pattern showed a new repeating structural unit at a spacing of 7.0 Å. Steaming and stretching the wool and then letting it shrink gave an x-ray pattern consistent with the original spacing of about 5.2 Å. Although these observations provided important clues to the molecular structure of wool, Astbury was unable to interpret them at the time.

a. What molecular properties of the wool fiber polypeptide support Atsbury’s observations?

b. Why does wool clothing shrink when washed in hot water or heated in a dryer whereas silk does not?

(a) The principal structural units in the wool fiber polypeptide, α-keratin, are α helices. Two α-keratin strands twisted into a coiled-coil produce the 5.2 Å spacing. Steaming and stretching the fiber yields an extended polypeptide chain with the β conformation in which the distance between adjacent R groups is about 7.0 Å. Upon re-steaming, the polypeptide chains again assume the less-extended α-helical conformation.

(b) Wool clothing shrinks when heated because it reverts to its native α-helical structure, whereas the very stable β-pleated sheets in silk remain unaffected. Freshly sheared wool is primarily in its α-helical coiled form. Because raw wool is crimped or curly, it is combed and stretched to straighten it before being spun into fibers for clothing. This processing converts the wool from its native α-helical conformation to a more extended β form. Moist heat triggers a conformational change back to the native α-helical structure, which shrinks both the fiber and the clothing

Proteinogenic amino acids, which are amino acids that are translated into proteins via the ribosome and can be chiral at the alpha carbon, are D-stereoisomers if they’re chiral at the alpha carbon

False

Biochemical processes are often coupled together to yield a net reaction with a DG greater than zero

False

Proteins with a higher Tm have a more positive DG of unfolding

True

A free energy diagram is a graph showing chemical potential on the y-axis and time on the x-axis

False

Below is the mechanism for the reaction catalyzed by ketosteroid isomerase; this is an example of covalent catalysis

False

A single hydrogen bond is shorter and stronger than van der Waal’s interactions discussed in this course

True

All molecules with at least one polar bond are polar molecules

False

All polar molecules have at least one polar bond

True

Proline is the only biological proteinogenic beta-amino acid

False

The resulting structure of a protein we determine an X-ray diffraction experiment is called an ensemble

False

A protein crystal structure solved to 1.5 angstroms is lower resolution than a crystal structure solved to 3.0 angstroms

False

The pKa of the carboxylic acid in glycine is lower than the carboxylic acid of acetic acid mostly due to stabilization by the nearby positively charged amine

True

The pKa of the amine of glycine is higher than the amine of methylamine mostly due to the inductive effects of the nearby deprotonated carboxylic acid (carboxylate)

False

Proteins composed of more than one peptide are described as having quaternary structure

True

Protein 3-D structures are more conserved than amino acid sequences

True

In a right-handed alpha helix composed of proteinogenic amino acids, the N-terminus of the helix has a partial negative charge, d-

False

When the pH of a solution is equal to the isoelectric (pI) of a solute molecule, the average solute molecule will have no charges present

False

Significantly greater than 50% of all amino acid residues in biological proteins are in the trans configuration about the peptide bond

True

Amide

Thioester

Thiol/thiolate/sulfhydryl

Phosphoryl

Aldehyde

Which of the following is a non-covalent interaction that only ionized functional groups are capable of participating in?

Salt bridges

Which of the following is NOT an axis label on the standard Ramachandran plot?

Omega

If the process A to B is spontaneous, which of the following statements must be true?

The DG of the process, as it is written, is less than zero

Which of the following describes the relationship of hydrogen bonds that stabilizes right-handed alpha helices that we discussed in class?

i+4

Which of the following describes the relationship of hydrogen bonds that stabilize beta sheets that we discussed in class?

There is no regular pattern to the network of hydrogen bonds that stabilize beta-sheets

Suppose a process A « B is coupled to process C « D; this results in the overall process A + C to B + D. The equilibrium constant of the process A « B is given by K1 = [B]eq/[A] eq, but the only equilibrium constant you can find for the process C « D is given by K2 = [C] eq /[D] eq. How would you combine these separate equilibrium constants to determine the equilibrium constant Kcoupled of the coupled process A + C to B + D?

Kcoupled = K1 * (1/K2)

We’ve seen three Ramachandran plots in MCB 354, and all three plots are shown below: the average amino acid, glycine, and proline. Assuming no other Ramachandran plots exist, and these are the only possible answers, which one corresponds to proline? Smiley face covers up y-axis label and is irrelevant to the answer.

Individual alpha-helices or beta-sheets are examples of what level of protein structure?

Secondary

Which of the following peptides is most likely to form a homodimeric helical bundle?

HLAENLREFLKNAR

Which of the following is NOT a model presented in class which helps explain how proteins limit conformational search space in order to fold in biologically relevant timeframes?

Induced Fit Model

Which of the following types of proteins generally uses energy from nucleotide triphosphates, such as ATP, to assist proteins that are misfolded in the folding process?

Chaperones

For the chemical process of A + B <-> P, k1 describes the rate of converting the substances on the left to product P. k-1 describes the rate of converting substance P to substances A and B. At equilibrium, which expression shows the relationship between the kinetic rate constants and the equilibrium constant describing the process above?

k1/k-1 = Keq

Catalysts decrease the energy of which species or value on a free energy diagram the most?

Transition state or states

Although all types of amino acids may appear in the interior of a globular protein, which general type of amino acid is most commonly found inside globular (otherwise known as cytoplasmic), folded proteins?

Non-polar

Which of the following terms describes the fundamental unit of protein tertiary structure that is a compact, self-contained, and folded structure that is independently stable in its native or near-native form?

Domain

When considering NMR couplings, which of these is not the same as the others?

Through-space coupling

Below is an image of a graph discussed in class labeled with Vmax and KM; which labels could be correct on the y-axis and x-axis? Answers listed as [insert y-axis], [insert x-axis]

Initial velocity, substrate concentration

Which molecular structure correctly shows the dominant charged species of peptide Met-His-Lys-Glu-Cys at pH 5.0?

A miniprotein was artificially designed and its stability assessed by thermal denaturation. The fraction of miniprotein unfolded at different temperatures was measured spectroscopically. What is the change in enthalpy (DHU°) for unfolding?

Temperature (C). Fraction Unfolded

40.0 0.207

45.0 0.407

47.0 0.500

50.0 0.636

160 kJ/mol

What is the change in entropy (DSU°) for unfolding of the designed miniprotein?

0.5 kJ/(mol*K)

What is the isoelectric point of the peptide RWLTGHASN?

10.4

The laboratory that you carry out research in has recently synthesized a new chromogenic substrate for the enzyme the lab studies. In order to use it in kinetic studies, you are tasked with determining the molar extinction coefficient of the molecule under the proposed assay conditions. You very carefully measure out a defined mass using the analytical balance and dissolve the mass in assay buffer, resulting in a solution of 1.17 mM concentration. If you use a standard 1 cm pathlength cuvette and measure an absorbance of 0.285, what is the molar extinction coefficient for this substance under this assay condition?

243.6 M-1cm-1

In gluconeogenesis, enolase catalyzes the conversion of phosphoenolpyruvate (PEP) to 2-phosphoglycerate (2-PG). The conversion of PEP to 2-PG has a DG°’ = -1.84 kJ/mol. What is the equilibrium constant? (Temperature = 37°C and R = 8.31 J mol-1 K-1)

2.042

In the absence of a catalyst, urea undergoes hydrolysis in water at a rate of 3 x 10-10 s-1. In the presence of urease, urea is hydrolyzed in water at a rate of 3 * 104 s-1. By what magnitude has the activation energy barrier been decreased in the presence of the catalyst? (Temperature = 25°C, R = 8.31 J mol-1 K-1)

80 kJ/mol

Alkaline phosphatase of sea shrimp (Pandalus borealis) crystallizes with unit cell dimensions a = 171.2 angstroms, b = 171.2 angstroms, and c = 84.3 angstroms. What is the angle of reflection, q, for the third order diffraction spot corresponding to Bragg planes parallel with the a-b plane and cutting through the c dimension (i.e. Miller indices [0 0 L])? The x-rays have wavelength 1.48 angstroms. Answers in degrees.

1.51

If the KD for an interaction is 10 nM (10*10-9 M), what is the DG of that interaction at 37 C? To clarify, this question is asking in reference to the following model or equation; the sign of the answer is part of the correct answer:

A + B to AB

-47.453 kJ/mol

Disulfide bonds found in proteins involve which amino acid(s)? Disulfide bonds are formed when two thiol group react with one another.

C only

Which of the following amino acids contains a sulfur atom? Choose all that apply.

M

Which of the following amino acids contain(s) a negatively charged group? Choose all that apply.

D and E

Of the 20 common amino acids, which amino acid(s) absorbs a significant amount of UV light? Choose all that apply.

Y and W

Which of the following groups of amino acids are most likely to be found clustered together in the hydrophobic core of a cytoplasmic protein (non-membrane protein)?

F, M, L, A, V

Which of the following amino acids contains an imidazole group?

H

Which of the following amino acids contains a guanidium group?

R

Monosodium glutamate (MSG) is a flavor enhancer often added to restaurant foods, canned vegetables, soups, deli meats and other foods. The U.S. Food and Drug Administration (FDA) has classified MSG as a food ingredient that's generally recognized as safe. Which amino acid is found in MSG?

E

Which amino acid is this?

None

Which amino acid is this?

Gly

Which of the following peptides is expected to dissolve readily in oil?

V-A-L-F-L-W

Which amino acid is this?

Ala

Which of the following groups of amino acids are most likely to be found clustered together on the surface of a protein cytoplasmic protein (non-membrane protein)?

R, N, Q, S, K

Which of the following cannot be spelled exclusively using the one letter code for the 20 common amino acids?

CELLO

Which of the following peptides is expected to dissolve readily in water?

M-Q-E-E-T

Which of the following amino acids contains an ionizable R group?

D

Which amino acid is this?

His

Which of the following statements about amino acids is/are true?

P contains an imino group

The primary structure of insulin is shown below. How many amino acids with a negatively charged side chain does insulin contain?

4

Q is an amide of which amino acid?

E

Which of the following amino acids is a non-chiral molecule?

G

Which amino acid is this?

None

Which of the following amino acids has the most hydrophilic R group?

E