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. 1983 Jan;80(2):344–348. doi: 10.1073/pnas.80.2.344

Detection and partial characterization of a chymostatin-sensitive endopeptidase in transformed fibroblasts.

J O'Donnell-Tormey, J P Quigley
PMCID: PMC393373  PMID: 6300832

Abstract

A chymostatin-sensitive step in the release of plasminogen activator from transformed fibroblasts has been described recently. By using synthetic peptidyl substrates, we have detected and characterized a chymostatin-sensitive peptidase activity in chicken embryo fibroblasts transformed by Rous sarcoma virus. The activity represents a neutral endopeptidase that exhibits phenylalanine specificity and is inhibited by diisopropyl fluorophosphate. A detailed inhibitor profile of the enzyme activity shows that it is distinct from other chymotrypsin-like phenylalanine-preferring peptidases. The endopeptidase activity in transformed fibroblasts is increased over that of parallel cultures of normal fibroblasts. The mechanism of enzyme inhibition by chymostatin is indicated by these studies, and the possible role of the enzyme in modulating plasminogen activator secretion is discussed.

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Selected References

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  1. Black P. H. Shedding from the cell surface of normal and cancer cells. Adv Cancer Res. 1980;32:75–199. doi: 10.1016/s0065-230x(08)60361-9. [DOI] [PubMed] [Google Scholar]
  2. Blobel G., Dobberstein B. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane-bound ribosomes of murine myeloma. J Cell Biol. 1975 Dec;67(3):835–851. doi: 10.1083/jcb.67.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. DIXON M. The effect of pH on the affinities of enzymes for substrates and inhibitors. Biochem J. 1953 Aug;55(1):161–170. doi: 10.1042/bj0550161. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dehm P., Prockop D. J. Time lag in the secretion of collagen by matrix-free tendon cells and inhibition of the secretory process by colchicine and vinblastine. Biochim Biophys Acta. 1972 Apr 21;264(2):375–382. doi: 10.1016/0304-4165(72)90302-9. [DOI] [PubMed] [Google Scholar]
  5. Diegelmann R. F., Peterkofsky B. Inhibition of collagen secretion from bone and cultured fibroblasts by microtubular disruptive drugs. Proc Natl Acad Sci U S A. 1972 Apr;69(4):892–896. doi: 10.1073/pnas.69.4.892. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Feinstein G., Janoff A. A rapid method for purification of human granulocyte cationic neutral proteases: purification and characterization of human granulocyte chymotrypsin-like enzyme. Biochim Biophys Acta. 1975 Oct 22;403(2):477–492. doi: 10.1016/0005-2744(75)90076-5. [DOI] [PubMed] [Google Scholar]
  7. Gerber A. C., Carson J. H., Hadorn B. Partial purification and characterization of a chymotrypsin-like enzyme from human neutrophil leucocytes. Biochim Biophys Acta. 1974 Sep 11;364(1):103–112. doi: 10.1016/0005-2744(74)90137-5. [DOI] [PubMed] [Google Scholar]
  8. Goldfarb R. H., Quigley J. P. Purification of plasminogen activator from Rous sarcoma virus transformed chick embryo fibroblasts treated with the tumor promoter phorbol 12-myristate 13-acetate. Biochemistry. 1980 Nov 25;19(24):5463–5471. doi: 10.1021/bi00565a001. [DOI] [PubMed] [Google Scholar]
  9. Jackson R. C., Blobel G. Post-translational cleavage of presecretory proteins with an extract of rough microsomes from dog pancreas containing signal peptidase activity. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5598–5602. doi: 10.1073/pnas.74.12.5598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  11. Lacy P. E., Howell S. L., Young D. A., Fink C. J. New hypothesis of insulin secretion. Nature. 1968 Sep 14;219(5159):1177–1179. doi: 10.1038/2191177a0. [DOI] [PubMed] [Google Scholar]
  12. Mumford R. A., Strauss A. W., Powers J. C., Pierzchala P. A., Nishino N., Zimmerman M. A zinc metalloendopeptidase associated with dog pancreatic membranes. J Biol Chem. 1980 Mar 25;255(6):2227–2230. [PubMed] [Google Scholar]
  13. O'Donnell-Tormey J., Quigley J. P. Inhibition of plasminogen activator release from transformed chicken fibroblasts by a protease inhibitor. Cell. 1981 Nov;27(1 Pt 2):85–95. doi: 10.1016/0092-8674(81)90363-9. [DOI] [PubMed] [Google Scholar]
  14. Pastan I., Almqvist S. Purification and properties of a mast cell protease. J Biol Chem. 1966 Nov 10;241(21):5090–5094. [PubMed] [Google Scholar]
  15. Powers J. C., Gupton B. F., Harley A. D., Nishino N., Whitley R. J. Specificity of porcine pancreatic elastase, human leukocyte elastase and cathepsin G. Inhibition with peptide chloromethyl ketones. Biochim Biophys Acta. 1977 Nov 23;485(1):156–166. doi: 10.1016/0005-2744(77)90203-0. [DOI] [PubMed] [Google Scholar]
  16. Quigley J. P. Association of a protease (plasminogen activator) with a specific membrane fraction isolated from transformed cells. J Cell Biol. 1976 Nov;71(2):472–486. doi: 10.1083/jcb.71.2.472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rindler-Ludwig R., Braunsteiner H. Cationic proteins from human neutrophil granulocytes. Evidence for their chymotrypsin-like properties. Biochim Biophys Acta. 1975 Feb 27;379(2):606–617. doi: 10.1016/0005-2795(75)90167-1. [DOI] [PubMed] [Google Scholar]
  18. Starkey P. M., Barrett A. J. Human cathepsin G. Catalytic and immunological properties. Biochem J. 1976 May 1;155(2):273–278. doi: 10.1042/bj1550273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Travis J., Bowen J., Baugh R. Human alpha-1-antichymotrypsin: interaction with chymotrypsin-like proteinases. Biochemistry. 1978 Dec 26;17(26):5651–5656. doi: 10.1021/bi00619a011. [DOI] [PubMed] [Google Scholar]
  20. Williams J. A., Wolff J. Possible role of microtubules in thyroid secretion. Proc Natl Acad Sci U S A. 1970 Dec;67(4):1901–1908. doi: 10.1073/pnas.67.4.1901. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Yurt R., Austen K. F. Preparative purification of the rat mast cell chymase: characterization and interaction with granule components. J Exp Med. 1977 Nov 1;146(5):1405–1419. doi: 10.1084/jem.146.5.1405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Zimmerman M., Ashe B. M. Sbustrate specificity of the elastase and the chymotrypsin-like enzyme of the human granulocyte. Biochim Biophys Acta. 1977 Jan 11;480(1):241–245. doi: 10.1016/0005-2744(77)90337-0. [DOI] [PubMed] [Google Scholar]
  23. Zimmerman M., Quigley J. P., Ashe B., Dorn C., Goldfarb R., Troll W. Direct fluorescent assay of urokinase and plasminogen activators of normal and malignant cells: kinetics and inhibitor profiles. Proc Natl Acad Sci U S A. 1978 Feb;75(2):750–753. doi: 10.1073/pnas.75.2.750. [DOI] [PMC free article] [PubMed] [Google Scholar]

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