Quantifying the fraction of alanine residues in an α-helical conformation in hornet silk using solid-state NMR

Here, we demonstrate the first successful isotope labeling of Ala carbons in hornet silk produced by the larvae of Vespa (Vespinae, Vespidae) mandarinia. This labeled hornet silk was examined by high-resolution C-13 solid-state NMR, and it was found that the fraction of Ala residues in alpha-helical conformations compared with Ala residues in the overall conformation of hornet silk can be quantitatively determined from Ala C-alpha NMR peaks. The value for this alpha-helical Ala fraction is close to that of the fraction of Ala residues in coiled-coil structures estimated in the four major hornet silk proteins by coiled-coil prediction analysis. This result indicates that most of the Ala residues in alpha-helices occur in those alpha-helices with a coiled-coil structure, and that the number of Ala residues in alpha-helices without a coiled-coil structure is small. Moreover, coiled-coil prediction analysis indicated that the potential coiled-coil domains are located only in the central portion of the protein chains of the major hornet silk proteins. From these results, we confirmed that the alpha-helical conformation mostly forms in the central portion of the hornet silk chains, whereas the ends of the protein chains are nearly devoid of alpha-helical structure. We deduce that the ends of the protein chains would preferentially adopt a beta-sheet conformation. Polymer Journal (2012) 44, 876-881; doi:10.1038/pj.2012.93; published online 23 May 2012