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. 1983 Jun 1;211(3):695–700. doi: 10.1042/bj2110695

Isolation and characterization of a trypsin-like serine proteinase from the membranes of Walker 256 carcino-sarcoma cells.

V J LaBombardi, E Shaw, J F DiStefano, G Beck, F Brown, S Zucker
PMCID: PMC1154416  PMID: 6349614

Abstract

A serine proteinase was isolated from Walker-256-carcino-sarcoma plasma-membrane-enriched preparations by affinity chromatography employing soya-bean trypsin inhibitor as the ligand. This enzyme was termed 'memsin' owing to its membrane location and trypsin-like substrate specificity. Analysis of this preparation by steric-exclusion high-pressure liquid chromatography (h.p.l.c.) resulted in a single peak of enzyme activity. Calculations of the rates of inactivation of memsin by peptidyl-chloromethanes and comparison with rate constants obtained with other serine proteinases indicated that memsin closely resembled trypsin and acrosin. Digestion of oxidized ribonuclease by memsin and analysis of the resulting peptides by h.p.l.c. yielded a chromatogram that was very similar to one generated by a tryptic digest of oxidized ribonuclease. This enzyme could possibly play a role in tumour-cell invasion.

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Selected References

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  1. Bosmann H. B., Bieber G. F., Brown A. E., Case K. R., Gersten D. M., Kimmerer T. W., Lione A. Biochemical parameters correlated with tumour cell implantation. Nature. 1973 Dec 21;246(5434):487–489. doi: 10.1038/246487a0. [DOI] [PubMed] [Google Scholar]
  2. Chrambach A., Reisfeld R. A., Wyckoff M., Zaccari J. A procedure for rapid and sensitive staining of protein fractionated by polyacrylamide gel electrophoresis. Anal Biochem. 1967 Jul;20(1):150–154. doi: 10.1016/0003-2697(67)90272-2. [DOI] [PubMed] [Google Scholar]
  3. Dewald B., Dulaney J. T., Touster O. Solubilization and polyacrylamide gel electrophoresis of membrane enzymes with detergents. Methods Enzymol. 1974;32:82–91. doi: 10.1016/0076-6879(74)32011-3. [DOI] [PubMed] [Google Scholar]
  4. DiStefano J. F., Beck G., Lane B., Zucker S. Role of tumor cell membrane-bound serine proteases in tumor-induced target cytolysis. Cancer Res. 1982 Jan;42(1):207–218. [PubMed] [Google Scholar]
  5. Green G. D., Shaw E. Thiobenzyl benzyloxycarbonyl-L-lysinate, substrate for a sensitive colorimetric assay for trypsin-like enzymes. Anal Biochem. 1979 Mar;93(2):223–226. doi: 10.1016/s0003-2697(79)80141-4. [DOI] [PubMed] [Google Scholar]
  6. Gurd J. W., Mahler H. R. Fractionation of synaptic plasma membrane glycoproteins by lectin affinity chromatography. Biochemistry. 1974 Dec 3;13(25):5193–5198. doi: 10.1021/bi00722a022. [DOI] [PubMed] [Google Scholar]
  7. Hatcher V. B., Wertheim M. S., Rhee C. Y., Tsien G., Burk P. G. Relationship between cell surface protease activity and doubling time in various normal and transformed cells. Biochim Biophys Acta. 1976 Dec 21;451(2):499–510. doi: 10.1016/0304-4165(76)90145-8. [DOI] [PubMed] [Google Scholar]
  8. Kettner C., Mirabelli C., Pierce J. V., Shaw E. Active site mapping of human and rat urinary kallikreins by peptidyl chloromethyl ketones. Arch Biochem Biophys. 1980 Jul;202(2):420–430. doi: 10.1016/0003-9861(80)90446-4. [DOI] [PubMed] [Google Scholar]
  9. Kettner C., Shaw E. Inactivation of trypsin-like enzymes with peptides of arginine chloromethyl ketone. Methods Enzymol. 1981;80(Pt 100):826–842. doi: 10.1016/s0076-6879(81)80065-1. [DOI] [PubMed] [Google Scholar]
  10. Kettner C., Shaw E. Synthesis of peptides of arginine chloromethyl ketone. Selective inactivation of human plasma kallikrein. Biochemistry. 1978 Oct 31;17(22):4778–4784. doi: 10.1021/bi00615a027. [DOI] [PubMed] [Google Scholar]
  11. Kettner C., Shaw E. The susceptibility of urokinase to affinity labeling by peptides of arginine chloromethyl ketone. Biochim Biophys Acta. 1979 Jul 11;569(1):31–40. doi: 10.1016/0005-2744(79)90078-0. [DOI] [PubMed] [Google Scholar]
  12. Kettner C., Springhorn S., Shaw E. Inactivation of boar acrosin by peptidyl-arginyl-chloromethanes. Comparison of the reactivity of acrosin, trypsin and thrombin. Hoppe Seylers Z Physiol Chem. 1978 Sep;359(9):1183–1191. doi: 10.1515/bchm2.1978.359.2.1183. [DOI] [PubMed] [Google Scholar]
  13. Liotta L. A., Tryggvason K., Garbisa S., Hart I., Foltz C. M., Shafie S. Metastatic potential correlates with enzymatic degradation of basement membrane collagen. Nature. 1980 Mar 6;284(5751):67–68. doi: 10.1038/284067a0. [DOI] [PubMed] [Google Scholar]
  14. Nachbar M. S., Oppenheim J. D., Aull F. Interactions of lectins with plasma membrane glycoproteins of the Ehrlich ascites carcinoma cell. Biochim Biophys Acta. 1976 Feb 6;419(3):512–529. doi: 10.1016/0005-2736(76)90262-5. [DOI] [PubMed] [Google Scholar]
  15. Polakoski K. L., McRorie R. A. Boar acrosin. II. Classification, inhibition, and specificity studies of a proteinase from sperm acrosomes. J Biol Chem. 1973 Dec 10;248(23):8183–8188. [PubMed] [Google Scholar]
  16. Rakonczay Z., Mallol J., Schenk H., Vincendon G., Zanetta J. P. Purification and properties of the membrane-bound acetylcholinesterase from adult rat brain. Biochim Biophys Acta. 1981 Jan 15;657(1):243–256. doi: 10.1016/0005-2744(81)90148-0. [DOI] [PubMed] [Google Scholar]
  17. Saklatvala J., Barrett A. J. Identification of proteinases in rheumatoid synovium. Detection of leukocyte elastase cathepsin G and another serine proteinase. Biochim Biophys Acta. 1980 Sep 9;615(1):167–177. doi: 10.1016/0005-2744(80)90020-0. [DOI] [PubMed] [Google Scholar]
  18. Unkeless J. C., Tobia A., Ossowski L., Quigley J. P., Rifkin D. B., Reich E. An enzymatic function associated with transformation of fibroblasts by oncogenic viruses. I. Chick embryo fibroblast cultures transformed by avian RNA tumor viruses. J Exp Med. 1973 Jan 1;137(1):85–111. doi: 10.1084/jem.137.1.85. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Zucker S., Lysik R. Cancer-induced cytolysis of normal bone marrow cells. Nature. 1977 Feb 24;265(5596):736–737. doi: 10.1038/265736a0. [DOI] [PubMed] [Google Scholar]

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