Distinct solid and solution state self-assembly pathways of RADA16-I designer peptide

Solid state NMR measurements on selectively C-13-labeled RADA16-I peptide (COCH3-RADARADARADARADA-NH2) were used to obtain new molecular level information on the conversion of -helices to -sheets through self-assembly in the solid state with increasing temperature. Isotopic labeling at the A4 C site enabled rapid detection of C-13 NMR signals. Heating to 344-363K with simultaneous NMR detection allowed production of samples with systematic variation of -helix and -strand content. These samples were then probed at room temperature for intermolecular C-13-C-13 nuclear dipolar couplings with the PITHIRDS-CT NMR experiment. The structural transition was also characterized by Fourier transform infrared spectroscopy and wide angle X-ray diffraction. Independence of PITHIRDS-CT decay shapes on overall -helical and -strand content infers that -strands are not observed without association with -sheets, indicating that -sheets are formed at elevated temperatures on a timescale that is fast relative to the NMR experiment. PITHIRDS-CT NMR data were compared with results of similar measurements on RADA16-I nanofibers produced by self-assembly in aqueous salt solution. We report that -sheets formed through self-assembly in the solid state have a structure that differs from those formed through self-assembly in the solution state. Specifically, solid state RADA16-I self-assembly produces in-register parallel -sheets, whereas nanofibers are composed of stacked parallel -sheets with registry shifts between adjacent -strands in each -sheet. These results provide evidence for environment-dependent self-assembly mechanisms for RADA16-I -sheets as well as new constraints on solid state self-assembled structures, which must be avoided to maximize solution solubility and nanofiber yields. Copyright (c) 2013 European Peptide Society and John Wiley & Sons, Ltd.