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. 1997 Feb;63(2):380–386. doi: 10.1128/aem.63.2.380-386.1997

Purification and characterization of two bifunctional chitinases/lysozymes extracellularly produced by Pseudomonas aeruginosa K-187 in a shrimp and crab shell powder medium.

S L Wang 1, W T Chang 1
PMCID: PMC168330  PMID: 9023918

Abstract

Two extracellular chitinases (FI and FII) were purified from the culture supernatant of Pseudomonas aeruginosa K-187. The molecular weights of FI and FII were 30,000 and 32,000, respectively, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 60,000 and 30,000, respectively, by gel filtration. The pIs for FI and FII were 5.2 and 4.8, respectively. The optimum pH, optimum temperature, pH stability, and thermal stability of FI were pH 8, 50 degrees C, pH 6 to 9, and 50 degrees C; those of FII were pH 7, 40 degrees C, pH 5 to 10, and 60 degrees C. The activities of both enzymes were activated by Cu2+; strongly inhibited by Mn2+, Mg2+, and Zn2+; and completely inhibited by glutathione, dithiothreitol, and 2-mercaptoethanol. Both chitinases showed lysozyme activity. The purified enzymes had antibacterial and cell lysis activities with many kinds of bacteria. This is the first report of a bifunctional chitinase/lysozyme from a prokaryote.

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Selected References

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  1. Chernin L., Ismailov Z., Haran S., Chet I. Chitinolytic Enterobacter agglomerans Antagonistic to Fungal Plant Pathogens. Appl Environ Microbiol. 1995 May;61(5):1720–1726. doi: 10.1128/aem.61.5.1720-1726.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Flach J., Pilet P. E., Jollès P. What's new in chitinase research? Experientia. 1992 Aug 15;48(8):701–716. doi: 10.1007/BF02124285. [DOI] [PubMed] [Google Scholar]
  3. Grinde B., Jollès J., Jollès P. Purification and characterization of two lysozymes from rainbow trout (Salmo gairdneri). Eur J Biochem. 1988 Apr 15;173(2):269–273. doi: 10.1111/j.1432-1033.1988.tb13994.x. [DOI] [PubMed] [Google Scholar]
  4. Gupta R., Saxena R. K., Chaturvedi P., Virdi J. S. Chitinase production by Streptomyces viridificans: its potential in fungal cell wall lysis. J Appl Bacteriol. 1995 Apr;78(4):378–383. doi: 10.1111/j.1365-2672.1995.tb03421.x. [DOI] [PubMed] [Google Scholar]
  5. 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]
  6. Legrand M., Kauffmann S., Geoffroy P., Fritig B. Biological function of pathogenesis-related proteins: Four tobacco pathogenesis-related proteins are chitinases. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6750–6754. doi: 10.1073/pnas.84.19.6750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Molano J., Polacheck I., Duran A., Cabib E. An endochitinase from wheat germ. Activity on nascent and preformed chitin. J Biol Chem. 1979 Jun 10;254(11):4901–4907. [PubMed] [Google Scholar]
  8. Monreal J., Reese E. T. The chitinase of Serratia marcescens. Can J Microbiol. 1969 Jul;15(7):689–696. doi: 10.1139/m69-122. [DOI] [PubMed] [Google Scholar]
  9. Ohta M., Yamagami T., Funatsu G. Purification and characterization of two chitinases from the leaves of pokeweed (Phytolacca americana). Biosci Biotechnol Biochem. 1995 Apr;59(4):656–661. doi: 10.1271/bbb.59.656. [DOI] [PubMed] [Google Scholar]
  10. Okazaki K., Kato F., Watanabe N., Yasuda S., Masui Y., Hayakawa S. Purification and properties of two chitinases from Streptomyces sp. J-13-3. Biosci Biotechnol Biochem. 1995 Aug;59(8):1586–1587. doi: 10.1271/bbb.59.1586. [DOI] [PubMed] [Google Scholar]
  11. Roby D., Gadelle A., Toppan A. Chitin oligosaccharides as elicitors of chitinase activity in melon plants. Biochem Biophys Res Commun. 1987 Mar 30;143(3):885–892. doi: 10.1016/0006-291x(87)90332-9. [DOI] [PubMed] [Google Scholar]
  12. Takayanagi T., Ajisaka K., Takiguchi Y., Shimahara K. Isolation and characterization of thermostable chitinases from Bacillus licheniformis X-7u. Biochim Biophys Acta. 1991 Jul 12;1078(3):404–410. doi: 10.1016/0167-4838(91)90163-t. [DOI] [PubMed] [Google Scholar]
  13. Ueda M., Fujiwara A., Kawaguchi T., Arai M. Purification and some properties of six chitinases from Aeromonas sp. no. 10S-24. Biosci Biotechnol Biochem. 1995 Nov;59(11):2162–2164. doi: 10.1271/bbb.59.2162. [DOI] [PubMed] [Google Scholar]
  14. Usui T., Hayashi Y., Nanjo F., Sakai K., Ishido Y. Transglycosylation reaction of a chitinase purified from Nocardia orientalis. Biochim Biophys Acta. 1987 Feb 20;923(2):302–309. doi: 10.1016/0304-4165(87)90017-1. [DOI] [PubMed] [Google Scholar]
  15. Usui T., Matsui H., Isobe K. Enzymic synthesis of useful chito-oligosaccharides utilizing transglycosylation by chitinolytic enzymes in a buffer containing ammonium sulfate. Carbohydr Res. 1990 Aug 1;203(1):65–77. doi: 10.1016/0008-6215(90)80046-6. [DOI] [PubMed] [Google Scholar]
  16. Wang S. L., Chang W. T., Lu M. C. Production of chitinase by Pseudomonas aeruginosa K-187 using shrimp and crab shell powder as a carbon source. Proc Natl Sci Counc Repub China B. 1995 Apr;19(2):105–112. [PubMed] [Google Scholar]
  17. Wang S. L., Pai C. S., Shieh S. T. Production of lytic enzyme from Pseudomonas aeruginosa M-1001. Proc Natl Sci Counc Repub China B. 1995 Oct;19(4):216–224. [PubMed] [Google Scholar]
  18. Wang S. L., Shieh S. T., Pai C. S. Production, purification and characterization of two proteinaceous hen-egg-white lysozyme inhibitors from Pseudomonas aeruginosa M-1001. Proc Natl Sci Counc Repub China B. 1995 Jul;19(3):166–175. [PubMed] [Google Scholar]
  19. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]

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