PH Dependence of the Coiled-Coil Structure of Keratin Intermediate Filament in Human Hair by 13C NMR Spectroscopy and the Mechanism of Its Disruption

Abstract

13C NMR spectra of the low sulfur fraction in S-(carboxymethyl)keratine (SCMKA) which corresponds to the hard-keratin intermediate filament (KIF) in human hair have been observed as a function of pH to clarify its disruption mechanism. The assignment was performed by the amino acid composition, both the distortionless enhancement by polarization transfer (DEPT) spectra and the chemical shift values of the SCMKA sample in 8 M urea solution. 13C NMR spectra at pH 5.0 and 6.0 contain essentially no peaks from the amino acid residues in the rod domain, which is due to its coiled-coil structure having highly restricted mobility. The coiled-coil structure was disrupted between pH 6.0 and 7.0 along with great great increase in peak intensities, which indicated the random coil structure occurred. During this disruption process, single chains with a helical form could not exist because there were no helical peaks in the spectra, or they existed for only a very short time, even if they did. Especially, the peak intensities of the side chains of the negatively charged amino acids, Glu and Asp, and those of the positively charged amino acids, Lys and Arg, increased abruptly at around pH 7.0, and these side chains formed ion-pairing interactions maintaining the coiled-coil structure in the rod domain. The peak intensities of the side chains of Leu and Ile also increased abruptly, indicating that hydrophobic interactions among these side chains in the coiled-coil structure were weakened. When the pH of the SCMKA solution was readjusted to pH 6.0 from 9.6, the 13C NMR spectrum was almost identical to that obtained originally at pH 6.0. Thus, both interactions are considered to contribute to the stability of the coiled-coil structure in the rod domain.