A CRITICAL CROSS-LINK REGION IN HUMAN-BONE-DERIVED COLLAGEN TYPE-I - SPECIFIC CLEAVAGE SITE AT RESIDUE LEU95

Collagen was extracted from human adult bone by limited pepsin digestion and collagen types were purified by consecutive salt precipitation first under neutral and then under acid conditions. In SDS/PAGE, all collagen type I preparations showed a protein band [α1s(I)] migrating between α(I) and α2(I) as well as a band [α2s(I)] migrating in front of α2(I). The collagenous nature of the pepsin‐stable α1s(I) protein was clearly demonstrated by digestion with human‐leucocyte‐derived collagenase, immunoblotting with antibodies against collagen type I and amino acid analysis. Partial amino acid sequencing of α1(I) and α1s(I) identified α1s(I) as a shortened α1(I) chain due to a specific cleavage site between residues Leu95 and Asp96 which is in close vicinity to the hydroxylysine‐derived crosslink at position 87. In circular dichroism, the proportion of thermally labile collagen molecules was proportional to the amount of shortened α1(I) and α2(I) chains, respectively. The melting temperature was found to be 36 ± 0.5°C as judged from circular dichroism and susceptibility to proteolysis. Our data provide clear evidence that a shortened α1‐derived collagen chain can be extracted from human adult bone whereas it is hardly found in human skin. The unique cleavage site might provide important information about the collagen I molecule embedded in the calcified matrix of human bone. Copyright © 1990, Wiley Blackwell. All rights reserved