Active Kallikrein, Preprokallikrein, and Kallikrein-Inhibitor Complex
Active kallikreins isolated from various exocrine and endocrine tissues were identified by a monoclonal antibody in Western blot analyses to be ~ 38,000 dalton proteins. Kallikreins isolated from rat pancreas, kidney, submandibular gland, brain, spleen and urine were indistinguishable with respect to molecular weight and immunological characteristics. Preprokallikreins were synthesized in a cell-free translation system directed by mRNAs and immunoprecipitated by affinity-purified kallikrein antibody. Analysis of the precipitates by SDS-polyacrylamide gel electrophoresis revealed a ~ 37,000 dalton polypeptide in kidney, brain and submandibular gland translation products. This 37,000 dalton kallikrein precursor was hybrid- arrested by a kallikrein cDNA encoding tissue kallikrein which was isolated from a rat submandibular gland cDNA library. The immunoprecipitates of products directed by pancreaticv mRNA showed a major protein with Mr of ~ 30,000. An endogenous ~ 92,000 dalton component in rat urine and kidney was also identified by a monoclonal antibody to tissue kallikrein and represents a kallikrein-inhibitor complex. These results indicate that tissue kallikreins can be initially synthesized as 37,000 or 30,000 dalton prepropeptides and then converted into a 38,000 dalton active form by proteolytic processing and glycosylation. The active kallikrein is capable of binding to an inhibitor to form a 92,000 dalton complex.
KeywordsTranslation Product Tissue Kallikrein Rabbit Reticulocyte Lysate Urinary Kallikrein Kidney Extract
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- 1.F. Fiedler, Enzymology of glandular kallikreins, in: “Handbook of Experimental Pharmacology,” Vol. 25, Supplement, Springer-Verlag, New York (1979).Google Scholar
- 2.J. J. Pisano, Chemistry and biology of the kallikrein-kinin system, in: “Proteases and Biological Control, Vol. 2, Cold Spring Harbor Conferences on Cell Proliferation,” E. Reich, D. B. Rifkin,, and E. Shaw, Cold Spring Harbor (1975).Google Scholar
- 3.J. Chao, C. Woodley, L. Chao, and H. S. Margolius, Identification of tissue kallikrein in brain and in the cell-free translation product encoded by brain mRNA, J. Biol. Chem., 58: 15173–15178 (1983).Google Scholar
- 4.J. Chao, L. Chao, and H. S. Margolius, Isolation of tissue kallikrein in rat spleen by monoclonal antibody-affinity chromatography, Biochem. Biophys. Acta, (In Press, 1984).Google Scholar
- 5.H. Nolly and M. C. Lama, Vascular kallikrein: A kallikrein-like enzyme present in vascular tissue of the rat, Clin. Sci., 63:249S–251S (1982).Google Scholar
- 7.J. Chao and H. S. Margolius, Isozymes of rat urinary kallikrein, Biochem. Pharmacol., 28: 2071–2079 (1979).Google Scholar
- 8.C. M. Woodley, J. Chao, H. S. Margolius, and L. Chao, Specific identification, stimulation or inhibition of rat tissue kallikrein with monoclonal antibodies, Kinin 84, Abstract, p. 150 (1984).Google Scholar
- 12.W. L. Gerald, J. Chao, and L. Chao, Sex dimorphism and hormonal regulation of rat tissue kallikrein mRNA (Manuscript Submitted, 1984 ).Google Scholar
- 13.W. L. Gerald, J. Chao, and L. Chao, isolation and analysis of a cDNA clone Encoding Rat Submaxillary Gland Kallikrein, DNA, 3: 86 (1984).Google Scholar