Radiochromatographic analysis of H2S-collagen complexes in rat-tail type I collagen.

Suspensions of 15 mg lyophilized type I, acid-soluble, rat-tail tendon collagen in 0.02 M tris/0.134 M NaCl (pH 7.4) buffer were reacted for up to 4 days at 4 degrees C with air containing [35S]-H2S. Gas-chromatography showed that all the H2S was absorbed: of the 46.8 per cent in the supernatant, 2.7 per cent was associated with neutral salt-soluble protein product and 53.2 per cent was complexed with a more cross-linked product which was soluble in 0.5 M acetic acid; approximately half (26.3 per cent of this product was stable in acid. The remaining 26.9 per cent was dissociated upon solubilization of the product in acetic acid and the cleaved fragment was separated by elution of paper chromatograms with 0.02 M tris/0.134 M NaCl buffer. The H2S-collagen complex was stable in this buffer, moderately stable in acetate/ethanol and highly unstable in an acetone/2-propanol/NH4OH solvent system. Thus there are at least two different binding sites on the type I collagen which react with H2S. One covalent bond, labile in acid, can be cleaved by dialysis of supernatant fraction in acetic acid. The second linkage, stable in acid within the time studied, is cleaved by acetone/2-propanol/NH4OH.