The effects of guanidine hydrochloride on the 'random coil' conformations and NMR chemical shifts of the peptide series GGXGG

The effects of the commonly used denaturant guanidine hydrochloride (GuHCl) on the random coil conformations and NMR chemical shifts of the proteogenic amino acids have been characterized using the peptide series Ac-Gly-Gly-X-Gly-Gly-NH2. The phi angle-sensitive coupling constants, ROESY cross peak intensities and proline cis-trans isomer ratios of a representative subset of these peptides are unaffected by GuHCl, which suggests that the denaturant does not significantly perturb intrinsic backbone conformational preferences. A set of (3)J(H)N(H) alpha values is presented which agree well with predictions of recently developed models of the random coil. We have also measured the chemical shifts of all 20 proteogenic amino acids in these peptides over a range of GuHCl concentrations. The shifts exhibit a linear dependence on denaturant concentration and we report here correction factors for the calculation of 'random coil' H-1 chemical shifts at any arbitrary denaturant concentration. Studies of a representative subset of peptides indicate that C-13 and N-15 chemical shifts are also perturbed by the denaturant. These results should facilitate the application of chemical shift-based analytical techniques to the study of polypeptides in solution with GuHCl. The effects of the denaturant on the quality of NMR spectra and on chemical shift referencing are also addressed.