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Chapter 06

1. a. The polyglycine helix falls under the polypeptide II helix structure and is a left-handed helix.
Text supporting Figures 6.5, 6.9, and Table 6.2.

b. See Table 6.1, 3.0 residues/turn.

2. (5 possible random disulfides) × (3 possible random disulfides) × (1 possible disulfide) = 15


possible random combinations of disulfides → 1 native combination/15 possible = 6.67% (1 in 15)
native disulfides would form randomly.

3. a. The four helices could be arranged so that the hydrophobic side chains would all point toward the
center of the bundle and would pack together there. This would give a stabilizing hydrophobic core.

b. A proline at this point would break the helix near the Fe 2 binding sites. This would probably
mean that Fe 2 could not be bound, and the mutant protein would be nonfunctional.

4. The α-helix is amphipathic. Each residue that is hydrophobic lies on one side of the helix due to the
repeated pattern, every 3–4 residues.

5. Pro is not able to adopt the ideal Φ and Ψ angles for an α-helix, nor does Pro have the α-NH group
that acts as a stabilizing H-bond donor in the middle of the helix.

6. a. 3200 = 2.7 × 1095

b. Not all of these conformations will be sterically possible. But even if only 0.1% of these are
allowed, there are still 2.7 × 1092 possible conformations—a very large number.

7.
a. ∆Sfolding = Sfolded − Sunfolded
= R ln Wfolded − R ln Wunfolded
= 8.314 J/K ⋅ mol ×[ln 1 − ln(2.7×1092 )]
= −1769 J/K ⋅ mol = −1.77 kJ/K ⋅ mol
b. ∆Hfolding = 96 × (−5 kJ/mol) = −480 kJ/mol
c. ∆Gfolding =∆Hfolding − T ∆Sfolding
= −480 kJ/mol − 298 K(−1.77 kJ/K ⋅ mol)
= +47 kJ/mol
Since ΔG folding > 0 at 25 °C, the protein would not be stable. It wold be stable below 0 °C, This points
out the importance of sources of stabilization other than backbone H bonds.

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Chapter 06

8. Output from the Jpred3 (http://www.compbio.dundee.ac.uk/www-jpred/index.html) server:


RRPVVLMAACLRPVVFITYGDGGTYYHWYH
---EEHHH-----EEEEEE-----EEEE--

From the PORTER server (http://distill.ucd.ie/porter/):


RRPVVLMAACLRPVVFITYGDGGTYYHWYH
CCCEEECCCCCCCEEEEEECCCCEEEEECC

These two secondary structure prediction programs (available via the ExPasy server) suggest that
there are three regions of beta-strand and the remainder of the peptide is “coil” (i.e., not helix or
sheet).

9. a. As in Figure 6.34, C 2 and D 2 both fit the requirement for a tetrameric protein with different
symmetry and interactions, the former being heterologous and the latter being isologous.

b. The highest symmetery possible is C 2 , because the quaternary structure of hemoglobin is a


dimer of α-β heterodimers (explained further in Chapter 7) and can thus only exhibit twofold
symmetry.

10. a. C 8 or D 4 .

b. D 4 , because it involves more subunit–subunit interactions.

c. Both. There must be heterologous interactions about the fourfold axis and isologous interactions
about the twofold axes.

11. The formation of favorable intramolecular ionic or H-bonding interactions in a folded protein
replace interactions between solvent (water) and the ionic species (or H-bond donors and acceptors) in
the unfolded state. The favorable ΔH obtained by formation of intramolecular bonds in the folded
protein is offset by the energy required to break many interactions, with solvent going from the
unfolded to the folded state.

12. a. MW = 18 + 115.09 + 163.18 + 147.18 + 129.12 + 114.11 + 103.14 + 97.12 + 128.18 + 57.06 =
1072.18g/mol. (Note: To the sum of residue masses, 1054.17 g/mol, you must add 18.01 g/mol for the
water equivalent that accounts for the free N-terminal amine and the C-terminal carboxylate in the
peptide.) From Figure 5.6, the molar absorptivity of Tyr is ~ 1000 M−1 cm−1. Since there is 1 Tyr per
molecule, this is also the molar absorptivity of vasopressin. This absorptivity can be converted to
units of cm2/mg as follows (recall that 1 L = 1000 mL and 1 mL = 1 cm3):

ε = 1000 M−1 cm−1 = 106 cm2 mol−1

ε = 106 cm2 mol−1 × (1 mol vasopressin)/1072.18 g) × (1 g/1000 mg) = 0.937 cm2 mg−1

b. A = εlc or 1.3 = (0.937 cm2 mg−1) × 0.5 cm × c

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Chapter 06

c = 2.8 mg/cm3

13. In the absence of BME, a single band of M W ≈ 70,000 is obtained. This suggests, but does not
prove, that there are two identical, noncovalently linked subunits. However, the addition of BME
removes this band and gives two bands of ≈ 30,000 and ≈ 40,000, respectively. The sum of these is
70,000, strongly suggesting that the native molecule contains four subunits, two of ≈ 30,000 and two
of ≈ 40,000. The 30,000 and 40,000 units are paired by at least one disulfide bond.

14. The fact that disulfide reduction has little effect means that the protein is a single chain. It must
have an extended structure, which cleavage at a critical Arg residue can relax, giving faster migration;
the fragments are still held together by disulfide bonds. Cleavage of these disulfides, after thrombin
cleavage, yields two fragments.

15. a. Assuming that X-ray diffraction is not practical, one could use CD, FT-IR, or NMR.

b. More than one secondary/tertiary folding can be observed for the same sequence. Therefore,
sequence alone cannot dictate folding in all cases, and sequence-based predictions must sometimes
fail.

16. a. No. DNA is charged and therefore polar, so most of the DNA-binding helix is likely to be
composed of polar residues that interact with either the DNA or the solvent. (This is supported by
sequence analysis of the bHLH family of DNA binding proteins.)

b. The N-terminus is interacting with the DNA. Two reasons might be given for this: (1) the
α-amino group of the N-terminus is positively charged and will interact favorably with the negative
charge on the phosphodiester backbone of the DNA; (2) this orientation also situates the “partial
positive” end of the helical macrodipole for favorable electrostatic interactions with the negatively
charged phosphodiester backbone of the DNA.

17. Consider how the mutation would affect the relative free energies in the folded and unfolded
states of the protein. Because the mutation is on the surface, loss of hydrophobic contacts in the
protein core is not an issue and the amino acid is likely to be interacting with solvent to similar
extents in both folded and unfolded states; thus, the effects on ΔH are predicted to be minimal. For the
same reason, the ΔS solvent is likely to be small because side-chain solvation is predicted to be similar in
both the folded and unfolded states (see Equation 6.3). ΔS protein changes the most due to the
conformational flexibility of Gly compared to Pro. Gly will stabilize both the folded and the unfolded
states; however, it stabilizes the unfolded state more due to the dramatic increase in conformational
entropy of the unfolded state as a result of this mutation. The stabilization of the unfolded state for the
mutant means that ΔG folding (wt) < ΔG folding(mutant) ; thus, the mutation is destabilizing.

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Chapter 06

18. The orientation of four contiguous residues is required to initiate one turn of α-helical structure
(Figure 6.4a). Likewise, a minimum of three contiguous residues must be ordered to initiate a turn
(Figure 6.20), which could then initiate H-bonding between antiparallel strands. Nucleation of an
antiparallel sheet can therefore be faster than nucleation of a helix. The initiation of a parallel sheet
requires a noncontiguous sequence to form H-bonding interactions. Because the effective
concentration of contiguous residues is higher than that of noncontiguous residues, the nucleation of a
parallel sheet will be significantly slower.

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