A High‐resolution 15 N Solid‐state NMR Study of Collagen and Related Polypeptides

Abstract

High‐resolution solid‐state 15 N‐NMR was used to clarify the effect of hydration on the stability of the coiled‐coil triple‐helix conformation of Gly‐X aa ‐Y aa repeating units in collagen and the collagen‐like polypeptides, (Pro‐Ala‐Gly) n , and (Pro‐Pro‐Gly) 10 , because the stability is thought to be related to the presence of (Gly)NH…O=C(X aa or Pro) hydrogen bonds. The 15 N‐NMR signals of these samples were narrowed upon hydration, mainly due to hydration‐induced conformational change or rearrangement of the repeating units. In particular, the 15 N chemical shifts of the Gly N‐H group and the high‐field (low‐frequency) shoulder peak of Pro nitrogen signals in (Pro‐Pro‐Gly) 10 were shifted downfield (4.9 ppm and 6.8 ppm, respectively) with increasing relative humidity, while the corresponding peaks for collagen and (Pro‐Ala‐Gly) n , were unchanged and close to the 15 N shift of (Pro‐Pro‐Gly) 10 , in the hydrated state. Such downfield shifts are consistent with the formation of N‐H…O=C hydrogen bonds. In agreement with the NMR results, it was found that the (Gly)NH…O=C (X aa or Pro) hydrogen bonds are retained in dehydrated collagen fibrils but not in partially dehydrated (Pro‐Pro‐Gly) 10 . No evidence was obtained for the partial formation of the 3 1 helix form (Pro) n or (Gly) n either under hydrated or dehydrated conditions. It is concluded that the Gly 15 N chemical shift is a very sensitive probe for studying supercoiling in collagen and collagen‐like polypeptides.