Analysis of chitinolytic enzyme genes of a mycoparasitic bacterium Flexibacter sp. FL824A
Description
[Synopsis]
We detected a lytic activity in a mycoparasite Flexibacter sp. FL824A toward several plant pathogenic fungi, consisting of the secretion of lytic enzymes, chitinolytic enzymes (chitinases and N-acetylglucosaminidases), β- 1,3-glucanases and proteases. Chitinolytic enzymes are considered to play an important role in the mycoparasitic process, because chitin, poly-β-1,4-N-acetyglucosamine, is a major structural component of cell walls of fungi.
A genomic library of Flexibacter sp. FL824A using pUC19 and Escherichia coli was constructed. Among 6,000 transformants tested, four clones showed a positive activity. They fell into two groups according to their specificity to the substrates. We selected two clones CHFll49 and CHF1351, based on the activity level, specificity to the substrates, and size of DNA-inserted fragment. An open reading frame of 4236 bp was found to code for a chitinase with 1412 amino acids based on the nucleotide sequence and deduced amino acid sequencing of an 8.7-kbp insert of pCHFI 149. Homology analysis of the deduced amino acid sequence of this protein revealed that the enzyme has a multiple domain structure consisting of at least seven domains. The most interesting property is that the enzyme has two catalytic domains, one homologous to the catalytic domain of chitinase A1 of Bacillus circulans WL-12 on the N-terminal side of the protein and the other homologous to that of chitinase D of the same bacterium on the C-terminal side. The DNA sequence of 5.0 kbp of the DNA-inserted fragment of pCHF1351 was determined after subcloning. An open reading frame of 1962 bp was found to code for a putative protein with 654 amino acids. The protein shared a 25% homology with endo-β-N-acetylglucosaminidase of Clostridiurn perfringens.
- Affiliation
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Japan International Research Center for Agricultural Sciences Animal Production and Grassland Division
- Classification
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Technical A
- Term of research
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FY1996-1998
- Responsible researcher
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ANDO Yasuo ( Animal Production and Grassland Division )
- ほか
- Japanese PDF
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1998_11_A3_ja.pdf417.38 KB