The Location of Cross-links in γ Chains from Calf Skin Collagen

Abstract

Abstract In calf skin collagen cross-links are formed between pairs of polypeptide chains and between all three chains of the tropocollagen molecules as the tissue matures. When the three chains are held in register by cross-links the molecule becomes reversibly renaturable. If such a molecule is transected by sonic irradiation or proteolytic attack then any part of the molecule that retains the cross-links between the three chains remains reversibly renaturable and can be fractionated and identified by electron microscopy. In this way we show that the cross-links occur at both ends of the molecule. The carboxyl-terminal quarter (B end) was identified as the major site of cross-links in all materials examined, but it was deduced that a Z chain structure (wherein a cross-link between two chains occurs at each end to one common chain) is the predominant species present in acid-soluble tropocollagen. Relatively few cross-links between all three chains occur in the amino-terminal quarter of acid-soluble tropocollagen and these are vulnerable to pepsin digestion; the cross-links at both ends were labile to Pronase digestion. In enzyme-solubilized collagen obtained from insoluble collagen (representing a more mature collagen), new or stabilized cross-links were found in both end quarters. Those at the amino terminus are resistant to pepsin or trypsin digestion, while those at the carboxyl terminus are resistant to pepsin, trypsin, and Pronase digestion. The possible existence of a structural weakness vulnerable to shearing forces at a point 43% of the length of the molecule from the carboxyl terminus and a structural discontinuity susceptible to trypsin action one-quarter length from the same end have been noted. The occurrence of a component higher in molecular weight than tropocollagen has been shown, and its possible relationship to a fundamental packing unit in fibrogenesis has been indicated.