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. 1997 Mar;63(3):834–839. doi: 10.1128/aem.63.3.834-839.1997

Molecular cloning, structural analysis, and expression in Escherichia coli of a chitinase gene from Enterobacter agglomerans.

L S Chernin 1, L De la Fuente 1, V Sobolev 1, S Haran 1, C E Vorgias 1, A B Oppenheim 1, I Chet 1
PMCID: PMC168378  PMID: 9055404

Abstract

The gene chiA, which codes for endochitinase, was cloned from a soilborne Enterobacter agglomerans. Its complete sequence was determined, and the deduced amino acid sequence of the enzyme designated Chia_Entag yielded an open reading frame coding for 562 amino acids of a 61-kDa precursor protein with a putative leader peptide at its N terminus. The nucleotide and polypeptide sequences of Chia_Entag showed 86.8 and 87.7% identity with the corresponding gene and enzyme, Chia_Serma, of Serratia marcescens, respectively. Homology modeling of Chia_Entag's three-dimensional structure demonstrated that most amino acid substitutions are at solvent-accessible sites. Escherichia coli JM109 carrying the E. agglomerans chiA gene produced and secreted Chia_Entag. The antifungal activity of the secreted endochitinase was demonstrated in vitro by inhibition of Fusarium oxysporum spore germination. The transformed strain inhibited Rhizoctonia solani growth on plates and the root rot disease caused by this fungus in cotton seedlings under greenhouse conditions.

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Selected References

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  1. Chen J. P., Nagayama F., Chang M. C. Cloning and expression of a chitinase gene from Aeromonas hydrophila in Escherichia coli. Appl Environ Microbiol. 1991 Aug;57(8):2426–2428. doi: 10.1128/aem.57.8.2426-2428.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Chinea G., Padron G., Hooft R. W., Sander C., Vriend G. The use of position-specific rotamers in model building by homology. Proteins. 1995 Nov;23(3):415–421. doi: 10.1002/prot.340230315. [DOI] [PubMed] [Google Scholar]
  4. De Filippis V., Sander C., Vriend G. Predicting local structural changes that result from point mutations. Protein Eng. 1994 Oct;7(10):1203–1208. doi: 10.1093/protein/7.10.1203. [DOI] [PubMed] [Google Scholar]
  5. Jones J. D., Grady K. L., Suslow T. V., Bedbrook J. R. Isolation and characterization of genes encoding two chitinase enzymes from Serratia marcescens. EMBO J. 1986 Mar;5(3):467–473. doi: 10.1002/j.1460-2075.1986.tb04235.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Lee B., Richards F. M. The interpretation of protein structures: estimation of static accessibility. J Mol Biol. 1971 Feb 14;55(3):379–400. doi: 10.1016/0022-2836(71)90324-x. [DOI] [PubMed] [Google Scholar]
  7. Perrakis A., Tews I., Dauter Z., Oppenheim A. B., Chet I., Wilson K. S., Vorgias C. E. Crystal structure of a bacterial chitinase at 2.3 A resolution. Structure. 1994 Dec 15;2(12):1169–1180. doi: 10.1016/s0969-2126(94)00119-7. [DOI] [PubMed] [Google Scholar]
  8. Rost B., Sander C. Combining evolutionary information and neural networks to predict protein secondary structure. Proteins. 1994 May;19(1):55–72. doi: 10.1002/prot.340190108. [DOI] [PubMed] [Google Scholar]
  9. Sitrit Y., Vorgias C. E., Chet I., Oppenheim A. B. Cloning and primary structure of the chiA gene from Aeromonas caviae. J Bacteriol. 1995 Jul;177(14):4187–4189. doi: 10.1128/jb.177.14.4187-4189.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Sobolev V., Wade R. C., Vriend G., Edelman M. Molecular docking using surface complementarity. Proteins. 1996 May;25(1):120–129. doi: 10.1002/(SICI)1097-0134(199605)25:1<120::AID-PROT10>3.0.CO;2-M. [DOI] [PubMed] [Google Scholar]
  11. Tsujibo H., Orikoshi H., Tanno H., Fujimoto K., Miyamoto K., Imada C., Okami Y., Inamori Y. Cloning, sequence, and expression of a chitinase gene from a marine bacterium, Altermonas sp. strain O-7. J Bacteriol. 1993 Jan;175(1):176–181. doi: 10.1128/jb.175.1.176-181.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Vriend G. WHAT IF: a molecular modeling and drug design program. J Mol Graph. 1990 Mar;8(1):52-6, 29. doi: 10.1016/0263-7855(90)80070-v. [DOI] [PubMed] [Google Scholar]
  13. Watanabe T., Kobori K., Miyashita K., Fujii T., Sakai H., Uchida M., Tanaka H. Identification of glutamic acid 204 and aspartic acid 200 in chitinase A1 of Bacillus circulans WL-12 as essential residues for chitinase activity. J Biol Chem. 1993 Sep 5;268(25):18567–18572. [PubMed] [Google Scholar]

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