Biotechnology and Bioprocess Engineering

, Volume 7, Issue 1, pp 38–42 | Cite as

Molecular cloning and characterization of 58 kDa chitinase gene fromSerratia marcescens KCTC 2172



A chitinase gene (pCHi58) encoding a 58 kDa chitinase was isolated from theSerratia marcescens KCTC 2172 cosmid library. The chitinase gene consisted of a 1686 bp open reading frame that encoded 562 amino acids.Escherichia coil harboring the pChi58 gene secreted a 58 kDa chitinase into the culture supernatant. The 58 kDa chitinase was purified using a chitin affinity column and mono-S column. A nucleotide andN-terminal amino acid sequence analysis showed that the 58 kDa chitinase had a leader peptide consisting of 23 amino acids which was cleaved prior to the 24th alanine. The 58 KDa chitinase exhibited a 98% similarity to that ofS. marcescens QMB 1466 in its nuclotide sequence. The chitinolytic patterns of the 58 kDa chitinase released N,N′-diacetyl chitobiose (NAG2) as the major hydrolysis end-product with a trace amount ofN-acetylglucosamine. When a 4-methylumbellyferyl-N-acetylglucosamin monomer, dimmer, and tetramer were used as substrates, the 58 kDa chitinase did not digest the 4-Mu-NAG monomer (analogue of NAG2), thereby indicating that the 58 kDa chitinase was likely an endochitinase. The optimum reaction temperature and pH of the enzyme were 50°C and 5.0, respectively.


Serratia marcescens chitinase endochitinase 


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  1. [1]
    Gooday, G. W., A. M. Humphreys, and W. H. McIntosh. (1986) Roles of chitinases in fungal growth In: R. Muzzarelli, C. Jeuniaux, and G. W. Gooday, (eds.).Chitin in Nature and Technology, pp. 83–91. Plenum Press, New York, USA.Google Scholar
  2. [2]
    Chtakara, A. (1961) Studies on the chitinolytic enzymes of black-koji mold: Purification of chitinase.Agric. Biol. Chem. 25: 54–60.Google Scholar
  3. [3]
    Boller, T. (1985) Induction of hydrolase as a defense reaction against pathogen, In: J. L. Key and T. Kosuge (eds.),Cellular and Molecular Biology of Plant Stress. pp. 247–262, A. R. Liss, New York, USA.Google Scholar
  4. [4]
    Powning, R. F. and H. Irzykiewez (1965) Studies on chitinase systems in bean and other seeds.Comp. Biochem. Physiol. 14: 127–133.CrossRefGoogle Scholar
  5. [5]
    Hamamura, Y. and Y. Kanehara (1940) Enzymatyic studies on exuvial fluid ofBombyx mori: Chitinase.J. Agric. Chem. Soc. Jpn. 16: 907–909.Google Scholar
  6. [6]
    Lunt, M. R. and P. W. Kent (1960) Chitinase system fromCarcinus maenas.Biochem Biophys. Acta 44: 371–373.CrossRefGoogle Scholar
  7. [7]
    Clarke, P. H. and M. V. Tracey (1956) The occurrence of chitinase in some bacteria.J. Gen. Microbiol. 14: 188–196.Google Scholar
  8. [8]
    Roberts, R. L. and E. Cabib (1982)Serratia marcescens chitinase: one-step purification and use for determination of chitin.Anal. Biochem. 127: 402–412.CrossRefGoogle Scholar
  9. [9]
    Watanabe, T., K. Suzuki, W. Oyanagi, K. Ohnishi, and H. Tanaka (1990) Gene cloning of chitinase A1 fromBacillus circulans WL-12 revealed its evolutionary relationship toSerratia chitinase and to the type III homology units of fibronectin.J. Biol. Chem. 265: 15659–15665.Google Scholar
  10. [10]
    Shinshi, H., D. Mohnen, and F. Meins (1987) Regulation of a plant pathogenesis-related enzyme: inhibition of chitinase and chitinase mRNA accumulation in cultured tabacco tissues by auxin and cytokinin.Proc. Natl. Acad. Sci. USA 84: 89–93.CrossRefGoogle Scholar
  11. [11]
    Fuchs, R. L., S. A. Mcpherson, and D. J. Drahos (1986) Cloning of aSerratia marcescens gene encoding chitinase.Appl. Environ. Microbiol. 51: 504–509.Google Scholar
  12. [12]
    Tsujibo, H., H. Orikoshi, K. Fujimoto, K. Miyamoto, C. Imada, Y. Okami, and Y. Inamori (1993) Cloning, sequence, and expression of a chitinase gene from a marine bacterium,Altermonas sp strain O-7.J. Bacteriol. 175: 178–181.Google Scholar
  13. [13]
    Wortman, A. T., C. C. Somerville, and R. R. Colwell (1986) Chitinase determinants ofVibrio vulnificus: Gene cloning and applications of a chitinase probe.Appl. Environ. Microbiol. 52: 142–145.Google Scholar
  14. [14]
    Sambrook, J., E. F. Fritsch, and T. Maniatis (1989)Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA.Google Scholar
  15. [15]
    Gal, S. W., J. Y. Choi, C. Y. Kim, Y. H. Cheong, Y. J. Choi, J. D. Bahk, S. Y. Lee, and M. J. Cho (1997) Isolation and characterization of the 54 kDa and 22 kDa chitinase genes ofSerratia marcescens KCTC 2172.FEMS Microbiol. Lett. 151: 197–204.Google Scholar
  16. [16]
    Gal, S. W., J. Y. Choi, C. Y. Kim, Y. H. Cheong, Y. J. Choi, J. D. Bahk, S. Y. Lee, and M. J. Cho (1998) Cloning of the 52-kDa chitinase gene fromSerratia marcescens KCTC 2172 and its proteolytic cleavage into an active 35-kDa enzyme.FEMS Microbiol. Lett. 160: 151–158.Google Scholar
  17. [17]
    Huber, M., E. Cabib, and L. H. Miller (1991)Proc. Natl. Acad. Sci. USA 88: 2807–2810.CrossRefGoogle Scholar
  18. [18]
    Vinetz, J. M., J. G. Valenzuela, C. A. Specht, L Aravind, R. C. Langer, J. M. C. Ribeiro, and D. C. Kaslow (2000) Chitinases of the Avian Malaria parasitePlasmodium gallinaceum, a class of enzymes necessary for parasite invasion of the mosquito midgut.J. Biol. Chem. 275: 10331–10341.CrossRefGoogle Scholar
  19. [19]
    Kless, H., Y. Sitrit, I. Chet, and A. B. Oppenheim (1989) Cloning of the gene coding for chitobias ofSerratia marcescens.Mol. Gen. Genet. 217: 471–473.CrossRefGoogle Scholar

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© The Korean Society for Biotechnology and Bioengineering 2002

Authors and Affiliations

  1. 1.Department of Microbiological EngineeringChinju National UniversityChinjuKorea
  2. 2.Department of Food and NutritionSila UniversityPusanKorea

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