Membrane-induced helical conformation of an active candidacidal fragment of salivary histatins.

The conformational preference of the candidacidal C-terminal 16 residue fragment (9-24; G-Y-K-R-K-F-H-E-K-H-H-S-H-R-G-Y) of salivary histatin 5 was examined in water, methanol, and dimethyl sulfoxide solutions using 500 MHz two-dimensional-NMR. Fourier transform infrared and CD spectroscopy were used to delineate its membrane-bound conformation in lipid vesicles. The peptide backbone and side-chain proton resonance assignments were accomplished by two-dimensional total correlated and nuclear Overhauser effect (NOE) spectra. The coupling constant (JNH-C alpha H) values determined from the double quantum-filtered correlated spectra, temperature coefficients of NH chemical shifts (d delta/dT), 1H/2H exchange rates on amide resonances, and the set of NOE connectivities were used to delineate backbone conformational features. The high JNH-C alpha H values (> or = 7.4 Hz), absence of any characteristic NH-NH (i, i + 1) or C alpha H-C beta H (i, i + 3) NOE connectivities, high d delta/dT values (> or = 0.004), and the fast 1H/2H amide exchange suggest that the histatin peptide favors unfolded random conformations in aqueous solution at pH 3.8. In contrast, the JNH-C alpha H values (< or = 6.5 Hz), slow 1H/2H exchange, low d delta/dT values (< or = 0.003) observed for amide resonances of residues 5-16, and the characteristic NH-NH (i, i + 1), C alpha H-C beta H (i, i + 3) NOE connectivities, provide evidence for the presence of largely alpha-helical conformations in dimethyl sulfoxide, which mimics the polar aprotic membrane environment. In methanolic solutions, 3(10)-helical conformations could exist as a minor population together with the major alpha-helical conformations. Fourier transform infrared spectroscopy and CD data indicate that lipid environments such as dimyristoylphosphatidylcholine vesicles could induce the peptide to fold into predominantly alpha-helical conformation. The results suggest that in dimethyl sulfoxide and dimyristoylphosphatidylcholine vesicles the candidacidal domain of salivary histatin 5 prefers a largely helical conformation, which could facilitate its interaction with the membrane of Candida albicans. The mechanism of antimicrobial action of this class of polypeptides appears to involve primarily electrostatic and hydrogen-bonding interaction of cationic and polar residues with the head groups of the plasma membranes of target cells.