Solution structures of TOAC-labeled trichogin GA IV peptides from allowed (g approximate to 2) and half-field electron spin resonance

The recently isolated trichogin GA IV is a 10 amino acid, Aib-rich peptide with potent membrane-modifying properties. The peptide is too short to span lipid bilayers, so the mechanism by which trichogin GA IV interacts with b iological membranes is unknown. The crystal structure has been solved, but there is much less information on the peptide's conformation in solution. T his problem is addressed by examining the electron spin resonance (ESR) of single and double TOAC-labeled trichogin GA IV analogues, where TOAC is a r igid nitroxide amino acid and serves as an Aib analogue. The doubly labeled peptides, trich-1,4, -4,8 and -1,8, represent all possible trichogin GA IV analogues containing two Aib --> TOAC substitutions. ESR in MeOH at 200 K of the g approximate to 2 spectral region suggests that the N-terminus from residues one through four adopts a helical structure similar to that obser ved in the crystal. However, the central and C-terminal regions appear to b e structurally heterogeneous. To further resolve the solution structure, we performed half-field ESR measurements in a MeOH/EtOH glass at 120 K and re ferenced them against similar measurements from a series of double TOAC-lab eled peptides of known structure. Half-field intensities depend on electron spin dipolar coupling and scale as 1/r(6) where r is the internitroxide di stance. The combination of allowed (g approximate to 2) and half-field ESR indicates that the trichogin GA IV C-terminal region is partially alpha-hel ical, as in the crystal structure, but is in equilibrium with unfolded conf ormers. It is suggested that the Gly-Gly stretch creates a hinge point betw een two short but stable helical regions. The combined ESR methods used her e represent a new approach for determining the solution structures of parti ally folded peptides.