The conformation of tridecapeptide alpha-mating factor from yeast Saccharomyces cerevisiae in aqueous solution was analyzed, in comparison with those of active analog and inactive analog peptides. 270-MHz 1H-NMR spectra of these peptides were observed and the spectral patterns of main-chain N-H proton resonances were classified into three groups. alpha-mating factor and Trp1-bearing active peptides belong to the group A1, active des-Trp1-peptides belong to the group A2 while the peptides of group B are inactive. The main-chain N-H proton resonances of the groups A1 and A2 and side-chain N-H proton resonances were all assigned to individual residues. The 13C-NMR analysis of alpha-mating factor indicates that the Lys7-Pro8 and Gln10-Pro11 peptide bonds exclusively take the trans form. From the temperature and pH dependences of chemical shifts and Gd(III)-induced relaxation enhancements of amide proton resonances, alpha-mating factor is found to take partly a folded conformation in aqueous solution, with an alpha-helical form in the N-terminal domain and two beta-turn forms in the central and C-terminal domain. The pH dependence of fluorescence intensity indicates that, in this folded conformation, the C-terminal carboxylate group lies close to the N-terminal domain. The presence of the folded form in the N-terminal domain and the beta-turn form in the central domain correlates with the biological activity of alpha-mating factor and analog peptides. However, the folded conformation of alpha-mating factor is in equilibrium with predominantly unordered form, as found from the circular dichroism and NMR analyses. The N-H proton and C-alpha proton resonances of free alpha-mating factor as assigned in the present study allow the transferred nuclear Overhauser enhancement (NOE) analysis of the membrane-bound conformation that is more directly related with the activity.

• PMID: 6323177
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Higashijima T, Masui Y, Chino N, Sakakibara S, Kita H, Miyazawa T

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