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. 1977 May;59(5):794–801. doi: 10.1172/JCI108701

Hydrolysis of the elastase substrate succinyltrialanine nitroanilide by a metal-dependent enzyme in rheumatoid synovial fluid.

J Saklatvala
PMCID: PMC372287  PMID: 16038

Abstract

A new type of enzyme hydrolyzing the elastase substrate succinyl-L-alanyl-L-alanine-4-nitroanilide has been found in cell-free rheuma todi synovial fluid. Normal plasma and osteoarthritic synovial fluid contained relatively little enzyme. The pH optimum was 8.0. Unexpectedly, the enzyme activity was not due to leukocyte elastase or any proteinase bound to alpha2-macroglobulin. The enzyme activity was metal-dependent being inhibited by chelating agents but not by di-isopropylfluorophos phate or thiol-blocking reagents. Gel chromatography showed the enzyme activity was associated with material of high molecular weight. On Sepharose 4B chromatography two-thirds of the activity eluted in the void volume and one-third in a position of about 106 mol wt. Utracentrifugation showed that both components were associated with lipid. The buoyant density of the higher molecular weight material was 1.15-1.22 g/ml., and that of lower molecular weight material was 1.2-1.33 g/ml. No latency of the enzyme was revealed by freezing and thawing or treatment with detergents. The nature of the enzyme is discussed. It is likely to be a proteinase possibly bound to some kind of membrane fragment.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barrett A. J., Starkey P. M. The interaction of alpha 2-macroglobulin with proteinases. Characteristics and specificity of the reaction, and a hypothesis concerning its molecular mechanism. Biochem J. 1973 Aug;133(4):709–724. doi: 10.1042/bj1330709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bieth J., Spiess B., Wermuth C. G. The synthesis and analytical use of a highly sensitive and convenient substrate of elastase. Biochem Med. 1974 Dec;11(4):350–357. doi: 10.1016/0006-2944(74)90134-3. [DOI] [PubMed] [Google Scholar]
  3. DINGLE J. T. Aetiological factors in the collagen diseases. Lysosomal enzymes and the degradation of cartilage matrix. Proc R Soc Med. 1962 Feb;55:109–111. [PubMed] [Google Scholar]
  4. ERLANGER B. F., KOKOWSKY N., COHEN W. The preparation and properties of two new chromogenic substrates of trypsin. Arch Biochem Biophys. 1961 Nov;95:271–278. doi: 10.1016/0003-9861(61)90145-x. [DOI] [PubMed] [Google Scholar]
  5. 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]
  6. Feinstein G., Janoff A. A rapid method of purification of human granulocyte cationic neutral proteases: purification and further characterization of human granulocyte elastase. Biochim Biophys Acta. 1975 Oct 22;403(2):493–505. doi: 10.1016/0005-2744(75)90077-7. [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. HAVEL R. J., EDER H. A., BRAGDON J. H. The distribution and chemical composition of ultracentrifugally separated lipoproteins in human serum. J Clin Invest. 1955 Sep;34(9):1345–1353. doi: 10.1172/JCI103182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Janoff A., Feinstein G., Malemud C. J., Elias J. M. Degradation of cartilage proteoglycan by human leukocyte granule neutral proteases--a model of joint injury. I. Penetration of enzyme into rabbit articular cartilage and release of 35SO4-labeled material from the tissue. J Clin Invest. 1976 Mar;57(3):615–624. doi: 10.1172/JCI108317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Janoff A., Scherer J. Mediators of inflammation in leukocyte lysosomes. IX. Elastinolytic activity in granules of human polymorphonuclear leukocytes. J Exp Med. 1968 Nov 1;128(5):1137–1155. doi: 10.1084/jem.128.5.1137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Keiser H., Greenwald R. A., Feinstein G., Janoff A. Degradation of cartilage proteoglycan by human leukocyte granule neutral proteases--a model of joint injury. II. Degradation of isolated bovine nasal cartilage proteoglycan. J Clin Invest. 1976 Mar;57(3):625–632. doi: 10.1172/JCI108318. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lazarus G. S., Daniels J. R., Brown R. S., Bladen H. A., Fullmer H. M. Degradation of collagen by a human granulocyte collagenolytic system. J Clin Invest. 1968 Dec;47(12):2622–2629. doi: 10.1172/JCI105945. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. MEHL J. W., O'CONNELL W., DEGROOT J. MACROGLOBULIN FROM HUMAN PLASMA WHICH FORMS AN ENZYMATICALLY ACTIVE COMPOUND WITH TRYPSIN. Science. 1964 Aug 21;145(3634):821–822. doi: 10.1126/science.145.3634.821. [DOI] [PubMed] [Google Scholar]
  14. Ohlsson K. Alpha1-antitrypsin and alpha2-macroglobulin. Interactions with human neutrophil collagenase and elastase. Ann N Y Acad Sci. 1975 Jun 13;256:409–419. doi: 10.1111/j.1749-6632.1975.tb36067.x. [DOI] [PubMed] [Google Scholar]
  15. Ohlsson K., Olsson I. The neutral proteases of human granulocytes. Isolation and partial characterization of two granulocyte collagenases. Eur J Biochem. 1973 Jul 16;36(2):473–481. doi: 10.1111/j.1432-1033.1973.tb02932.x. [DOI] [PubMed] [Google Scholar]
  16. Pryce-Jones R. H., Saklatvala J., Wood G. C. Neutral protease from the polymorphonuclear leucocytes of human rheumatoid synovial fluid. Clin Sci Mol Med. 1974 Nov;47(5):403–414. doi: 10.1042/cs0470403. [DOI] [PubMed] [Google Scholar]
  17. Pryce-Jones R. H., Wood G. C. Purification of granulocyte neutral protease from human blood and rheumatoid synovial fluid. Biochim Biophys Acta. 1975 Aug 26;397(2):449–458. doi: 10.1016/0005-2744(75)90135-7. [DOI] [PubMed] [Google Scholar]
  18. Saklatvala J., Wood G. C., White D. D. Isolation and characterization of human plasma alpha 1-proteinase inhibitor and a conformational study of its interaction with proteinases. Biochem J. 1976 Aug 1;157(2):339–351. doi: 10.1042/bj1570339. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Weissmann G. Lysosomal mechanisms of tissue injury in arthritis. N Engl J Med. 1972 Jan 20;286(3):141–147. doi: 10.1056/NEJM197201202860307. [DOI] [PubMed] [Google Scholar]

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