ESR characterization of hexameric, helical peptides using double TOAC spin labeling

alpha-Aminoisobutyric acid (Aib) is a C-alpha-tetrasubstituted amino acid that strongly favors helical structure. Most of the conformational trends established for Aib-rich peptides have been determined by X-ray crystallography. Whether these conformational trends carry over to protic solvents is an open question. In order to develop a general strategy for probing the properties of peptides containing C-alpha-tetrasubstituted amino acids, the hexameric sequences Boc-TOAC-Ala(n)-TOAC-Ala(4-n)-OtBu were synthesized where n = 0-3 and TOAC is a spin label Aib analog, The peptides were studied by electron spin resonance (ESR) in four alcohols: MeOH, EtOH, TFE, and HFIP. Biradical J-coupling and dipolar interactions between the TOACs within each peptide were used to determine peptide geometry as a function of solvent. In MeOH, strong biradical interactions were observed consistent with the geometry of a 3(10)-helix. The solvents displayed differing tendencies to support helical structure with the ranking MeOH > EtOH > TFE > HFIP. In HFIP, there were no indications of residual helical structure. While C-alpha-tetrasubstituted amino acids do favor the helix, these data demonstrate that such amino acids do not ''lock in'' the helical conformation. Qualitative analysis of the line width variations for the hexamers in MeOH suggests that the interconversion time for helix --> coil is several nanoseconds. Additional peptides were prepared in order to explore the effects of peptide length, N-terminal blocking group, and insertion of an additional Aib.