VCD spectroscopic properties of the β‐hairpin forming miniprotein CLN025 in various solvents

Abstract

Electronic and vibrational circular dichroism are often used to determine the secondary structure of proteins, because each secondary structure has a unique spectrum. In order to determine these spectral features, polypeptides that are known to adopt a particular conformation along their entire length are studied ideally. Little is known about the vibrational circular dichroic spectroscopic features of the β-hairpin. In this study, the VCD spectral features of a decapeptide, YYDPETGTWY (CLN025), which forms a stable β-hairpin that is stabilized by intramolecular weakly polar interactions and hydrogen bonds were determined. Molecular Dynamics simulations and ECD spectropolarimetry were used to confirm that CLN025 adopts a β-hairpin in water, TFE, MeOH and DMSO and to examine differences in the secondary structure, hydrogen bonds and weakly polar interactions. CLN025 was synthesized by microwave-assisted solid phase peptide synthesis with Nα-Fmoc protected amino acids. The VCD spectra displayed a (−,+,−) pattern with bands at 1640 to 1656 cm−1, 1667 to 1687 cm−1 and 1679 to 1686 cm−1 formed by the overlap of a lower frequency negative couplet and a higher frequency positive couplet. A maximum IR absorbance was observed at 1647 to 1663 cm−1 with component bands at 1630 cm−1, 1646 cm−1, 1658 cm−1 and 1675 to 1680 cm−1 that are indicative of the β-sheet, random meander, either random meander or loop and turn, respectively. These results are similar to the results of others, who examined the VCD spectra of β-hairpins formed by DPro-Xxx turns and indicate that observed pattern is typical of β-hairpins.

Aryl–aryl interactions in designed peptide folds: Spectroscopic characteristics and optimal placement for structure stabilization

“…For many of the clusters the 195nm exciton couplet is the dominant CD feature. We found no evidence of the −[θ] 196 /+[θ] 202 exciton couplet predicted by Roy and Keiderling 27,28 for the Y E /W F cluster and attributed to a Y/Y interaction by Lovas 30,32 . We also have seen no evidence for a significant CD contribution from diagonal face-to-face interaction W/W interactions in hairpins.…”

“…5 provides a comparison of the CD spectra reported for - W F T Y E -ENGK- W F T Y E -, chignolin and CLN025. In the case of CLN025, quite different CD spectra were reported by the Sandor Lovas group 30–32 in an extensive investigation of the effects of organic solvents and chemical denaturants which was supported by MD simulations.…”

“…The CD changes associated with the added tyrosines are exclusively at λ < 210 nm, most notably a maximum at 203 nm. In his discussion of CLN025 CD spectra, Lovas 30,32 viewed the extrema at 197 and 203 nm as another exciton couplet, but did not attribute it to a specific interaction. Keiderling noted 27 that W F /Y E interactions were expected 28 to produce a −[θ] 195 /+[θ] 200 exciton couplet feature and indicated that the dominant −[θ] 212 /+[θ] 228 feature of W F Y E -- W F Y E -TZ2 was a W/W exciton couplet, presumably associated with the diagonal stack-shifted face-to-face interaction of the W2 and W9.…”

Molecular Dynamics Analysis of the Conformations of a β-Hairpin Miniprotein

“…The backbone RMSD values between the middle structures from the simulations and the X-ray structure are lower than the RMSD value of the first NMR structure and only the RMSD values between the backbone of the peptide at 300 K and 363 K (simulations 3, 4 ) and the NMR structure are greater than the RMSD value of the X-ray structure. ECD and VCD spectra of CLN02544 confirm that the peptide maintains a β-hairpin in the environments studied and that in TFE and MeOH the turn of the peptide changes.…”

The CLN025 Decapeptide Retains a β-Hairpin Conformation in Urea and Guanidinium Chloride

“…30,31 In this study, MD simulations and Electronic Circular Dichroism (ECD) spectropolarimetry are used to investigate the conformational stability of the CLN025 β-hairpin in different concentrations of urea and GdmCl and to examine the changes in the H-bonds and weakly polar interactions. Conformational stability was investigated by ECD using 0 to 6 M GdmCl and 0 to 8 M urea in 0.5 M increments.…”