Organic Letters
Letter
residues, i.e., residues i and i + 3, stabilize this expanded β-turn.
The fact that glycine and β-alanine, the least conformationally
restricted α- and β-amino acids, are accommodated together in
this expanded β-turn suggests that replacing the glycine and/or
β-alanine residue(s) with other α- and β-amino acid residues
that are conformationally more restricted could result in
analogous β-turns with varied and perhaps enhanced stabilities.
Our initial studies indicated that replacing the glycine and β-
alanine residues of 1 with L-alanine and L-β-homoalanine,
respectively, led to tetrapeptides that also adopt hairpin
conformations. This result suggests that the expanded β-turn
formed by 1 may represent a general turn motif capable of
accommodating different α- and β-amino acid residues.
Herein we report the effects of replacing the glycine and β-
alanine residues of 1 with α-amino acid and L-β-homoamino
acid residues on the folding of the resultant hybrid peptides
and the stabilities of the corresponding hairpins. Hybrid
peptide 2a differs from 1 only in its side chains (R groups).
Peptides 2b−j result from replacing the glycine and β-alanine
residues of 2a with different α- and β-amino acid residues
Table 1. Differences in the Chemical Shifts of the Amide
Protons of 2a−j at Low and High Concentrations
a
b
ΔδNH (ppm)
entry
a
b
c
d
e
2a
2b
2c
2d
2e
2f
−0.048
−0.129
−0.120
−0.127
−0.018
−0.055
−0.154
−0.008
−0.082
−0.054
−0.045
−0.051
−0.027
−0.099
0.061
−0.036
−0.075
0.027
0.854
0.845
0.664
0.422
0.485
0.222
0.593
0.073
0.187
0.825
−0.032
0.086
0.117
0.236
0.077
0.011
0.050
0.003
0.022
0.132
−0.046
−0.048
−0.060
−0.046
−0.047
−0.023
−0.073
−0.015
0.042
1
7
2g
2h
c
2i
−0.020
−0.118
2
j
−0.155
a
1
H NMR spectra were recorded in CDCl (400 MHz, 298 K).
3
b
c
ΔδNH = δ(25 mM) − δ(0.1 mM). ΔδNH = δ(1.0 mM) − δ(0.1 mM)
.
In mixed solvents with different ratios of DMSO-d and
6
CDCl , the amide proton resonances of 2a−j show shifts that
3
are consistent with the adoption of hairpin conformations
(
Table 2). From 0 to 40% DMSO-d , among the signals of the
6
Table 2. Differences in the Chemical Shifts of Amide
a
Protons in CDCl Containing 0 and 40% DMSO-d6
3
b
ΔδNH (ppm)
entry
a
b
c
d
e
2
2
2
2
2
a
b
c
d
e
0.067
0.311
0.239
0.428
0.343
−0.013
0.049
−0.052
−0.040
0.091
−0.288
−0.230
−0.297
−0.254
−0.280
−0.136
−0.039
−0.240
−0.142
−0.049
1.041
1.217
1.444
0.889
0.970
1.558
1.705
1.705
1.885
1.577
0.404
0.418
0.738
0.553
0.347
0.225
0.323
0.197
0.616
0.510
−0.121
−0.135
−0.142
−0.135
−0.137
−0.120
−0.115
−0.116
−0.195
−0.100
Figure 2. Hybrid peptides 2a−j shown as their likely hairpin
conformation.
2f
2g
c
2
2
2
h
i
j
Our study shows that substituting the glycine residue of 2a
with four L-α-amino acid residues, Ala, Phe, Val, and Met, gives
hybrid peptides 2b, 2c, 2d, and 2e that fold into hairpins with
stabilities similar to or slightly lower than that of 2a; replacing
the β-alanine residue of 1 with three L-β-homoamino acids, β-
hAla, β-hPhe, and β-hVal, results in peptides 2f, 2g, and 2h
that fold into hairpins with enhanced stabilities. In addition,
hybrid peptides 2i and 2j, which both have the L-β-hAla
residue but differ in their L- and D-α-Phe residues, respectively,
fold into hairpins with different stabilities. The obtained results
demonstrate that the expanded β-turn observed with 1 is a
general turn motif capable of accommodating α- and β-amino
acid residues bearing different side chains.
a
Proton NMR spectra were recorded in CDCl at 5 mM (600 MHz,
3
b
c
298 K). ΔδNH = δ(40% DMSO) − δ(0% DMSO). Spectra were measured at
1
mM because of limited solubility of 2h in CDCl .
3
five amide protons of each peptide, that of proton c shows the
largest shift, from ∼1 ppm to nearly 2 ppm, indicating that
proton c is exposed to solvent molecules and becomes
increasingly H-bonded as more polar (i.e., DMSO-d6)
molecules become available. In contrast, the resonances of
the other four amide protons show negligible or much smaller
shifts, from less than 0.1 ppm to about 0.6 ppm, consistent
with their involvement in intramolecular H-bonding.
1
H NMR spectra of 2a−j recorded from 0.1 to 25 mM (from
Changes in the chemical shifts of protons a and d of 2a−g,
0
.1 to 1 mM for 2h because of limited solubility) in CDCl
2i, and 2j in CDCl from 0 to 45 °C, which indicate the
3
3
revealed that the amide proton resonances of peptides 2b−j
follow the same trend shown by 2a. Among the resonances of
the five amide protons of each hybrid peptide, that of proton c
shows the largest shift (Table 1). Similar to the signals of
intramolecularly H-bonded protons b and e, the resonances of
protons a and d exhibit either insignificant or small shifts with
increasing concentration. These observations are consistent
with the adoption of the hairpin conformation (Figure 2)
found with 1 (or 2a) by each of these peptides, in which
protons a and d are intramolecularly H-bonded while proton c
engages in intermolecular H-bonding.
stabilities of the H-bonds involving these protons, were
compared to qualitatively estimate the stabilities of the hairpin
conformations. The resonances of protons a of 2b, 2c, 2d, and
2e show upfield shifts similar to that observed for proton a of
2a, indicating that substituting the glycine residue of 2a with
other α-amino acid residues does alter the strength of the
intramolecular H-bonds involving protons a (Figure 3a, blue
columns). In contrast, the shifts observed for the signals of
protons a of 2f and 2g are noticeably smaller than that shown
by the resonance of proton a of 2a, suggesting that substituting
the β-alanine residue of 2a with β-amino acid residues bearing
B
Org. Lett. XXXX, XXX, XXX−XXX