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 (G uHCl) on the random coil conformations and NMR chemical shifts of the proteogenic amino acids have been characterized using the peptide seri es Ac-Gly-Gly-X-Gly-Gly-NH2. The phi angle-sensitive coupling constant s, ROESY cross peak intensities and proline cis-trans isomer ratios of a representative subset of these peptides are unaffected by GuHCl, wh ich suggests that the denaturant does not significantly perturb intrin sic backbone conformational preferences. A set of (3)J(H)N(H) alpha va lues is presented which agree well with predictions of recently develo ped models of the random coil. We have also measured the chemical shif ts of all 20 proteogenic amino acids in these peptides over a range of GuHCl concentrations. The shifts exhibit a linear dependence on denat urant concentration and we report here correction factors for the calc ulation of 'random coil' H-1 chemical shifts at any arbitrary denatura nt concentration. Studies of a representative subset of peptides indic ate that C-13 and N-15 chemical shifts are also perturbed by the denat urant. These results should facilitate the application of chemical shi ft-based analytical techniques to the study of polypeptides in solutio n with GuHCl. The effects of the denaturant on the quality of NMR spec tra and on chemical shift referencing are also addressed.