Free metal ion-independent oxidative damage of collagen. Protection by ascorbic acid

In this paper we demonstrate that in the absence of free metal ions, active oxygen species, generated by activated macrophages or xanthine/xanthine oxidase (XOD), carry out oxidative degradation of collagen fibrils type I in conjunction with proteases. The collagen degradation is completely prevented by ascorbate (AH2) but not by catalase. The free metal ion-independent collagen degradation is a two-step process: (i) oxidation of collagen and (ii) subsequent proteolytic cleavage of the oxidatively modified collagen. AH2 completely prevents collagen oxidation and thereby protects the collagen from subsequent proteolytic degradation. This is in contrast to free metal ion-catalyzed spontaneous fragmentation of collagen, which is accelerated by AH2 and inhibited by catalase (Kato, Y., Uchida, K., and Kawakishi, S. (1992) J. Biol. Chem. 267, 23646-23651). Studies using xanthine/XOD and model polypeptides, namely, poly-L-Pro, poly-L-hydroxyproline, poly-L-Lys, and poly(Pro-Gly-Pro) indicate that although O2-. is needed along with XOD, oxidation of model polypeptides appears to be a direct function of XOD iron, which is also stimulated by cytochrome P450.