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 top 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 was also found to cleave the C-propeptides from type II and type III procollagens.
Human epidermal growth factor (EGF) was acylated at its three primary amine groups. The acylated derivatives were purified, characterized, and their activities and response to trypsin were compared with unmodified EGF. Modified EGF was shown to retain its biological activity but displayed a higher shelf stability and resistance to tryptic digestion in aqueous solutions as compared with the unmodified growth factor.
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