Isolation and partial characterization of kinin-like peptides formed by acid protease from murine fibroblast L-929.

A kinin-forming acid protease was isolated and purified from a stationary cell culture of murine fibroblasts L-929 (Biochem. Pharmacol. 26: 1187, 1977). The enzyme formed kinins at an optimal pH of 4.0 from Murphy-Sturm lymphosarcoma tissue and rat plasma kininogen substrate. This study reports on the isolation and partial chemical characterization of two fibroblast kinins. The 12,000 g fraction of L-929 fibroblast homogenates was incubated with rat plasma at 37 degrees, pH 4.0, for 92 hours, and the kinins extracted with ethyl alcohol-p-toluene sulfonic acid. Two fibroblast kinins, FKI and FKII, were separated and purified on the following chromatographic columns: G-25 Sephadex (1 X 115 cm), CM-C50 Sephadex (2.5 X 15 cm), and Biogel P-4 (1 X 115 cm). Kinin activity was bioassayed on the perfused isolated rat uterus preparation. The purity of the kinins was ascertained by thin layer chromatography coupled with dansylation. The estimated molecular weight of FKI was 1450 with a 14-amino acid composition (determined by automatic AA analysis) that included AspThrSerProGluGlyAlaValLeuTyrPheLysHisArg. FKII had an estimated molecular weight of 1000 with a 12-amino acid composition including AspSerProGluGlyAlaLeuTyrLysHisArg. The kinins were relatively rich in arginine and did not contain the bradykinin sequence.