Partial purification of a procollagen C-proteinase. Inhibition by synthetic peptides and sequential cleavage of type I procollagen

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Abstract
A procollagen C-proteinase which cleaves the C-propeptides from type I procollagen was purified about 125-fold from membranous bones of chick embryos. As estimated by gel filtration, the enzyme was about 80,000 daltons. When a reaction with modified procollagen was carried out, the enzyme preferentially cleaved the C-propeptides from the pro.alpha. chains in the order pro.alpha.1, pro.alpha.1 and then pro.alpha.2. The enzyme was inhibited by several metal chelators and high concentrations of dithiothreitol. It was also inhibited by 5% fetal calf serum. A series of inhibitors of serine proteinases and sulfhydryl-containing proteinases were not inhibitory. Four oligopeptides were synthesized with amino acid sequences, similar to the amino acid sequences around the sites at which the C-propeptides are cleaved during the conversion of procollagen to collagen in vivo. The peptide Tyr-Tyr-Arg-Ala-Asp-Asp-Ala inhibited the enzyme 35-60% in concentrations of 6-12 mM. Shorter peptides containing the Ala-Asp bond cleaved by the enzyme were less effective. The partially purified enzyme cleaved the C-propeptides from type II and type III procollagens.