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MOLECULAR CLONING OF CHITINASE 33 (CHIT33) GENE FROM TRICHODERMA ATROVIRIDE

In this study Trichoderma atroviride was selected as over producer of chitinase enzyme among 30 different isolates of Trichoderma sp. on the basis of chitinase specific activity. From this isolate the genomic and cDNA clones encoding chit33 have been isolated and sequenced. | Brazilian Journal of Microbiology 2008 39 433-437 ISSN 1517-8382 MOLECULAR CLONING OF CHITINASE 33 CHIT33 GENE FROM TRICHODERMA ATROVIRIDE Matroudi S. Zamani M.R. Motallebi M. National Institute for Genetic Engineering and Biotechnology NIGEB Tehran I.R. of Iran Submitted July 17 2007 Returned to authors for corrections November 22 2007 Approved July 06 2008. ABSTRACT In this study Trichoderma atroviride was selected as over producer of chitinase enzyme among 30 different isolates of Trichoderma sp. on the basis of chitinase specific activity. From this isolate the genomic and cDNA clones encoding chit33 have been isolated and sequenced. Comparison of genomic and cDNA sequences for defining gene structure indicates that this gene contains three short introns and also an open reading frame coding for a protein of 321 amino acids. The deduced amino acid sequence includes a 19 aa putative signal peptide. Homology between this sequence and other reported Trichoderma Chit33 proteins are discussed. The coding sequence of chit33 gene was cloned in pEt26b expression vector and expressed in E. coli. Key-words Trichoderma atroviride chit33 chitinase activity gene structure. Chitin is a polymer of P-1 4 linked N-acetylglucosamine GlcNAc and a very abundant natural polymer. It is the main structural compound of cell wall of fungi insect exoskeletons and shells of crustaceans 10 . The fungal cell wall is a highly dynamic structure subject to constant change during cell expansion and division and during spore germination hyphal branching and septum formation in filamentous fungi. The cell wall degrading enzymes are glycosyl hydrolases that degrade chitin and glucan polymer which comprise important structural elements in the cell walls of fungal organisms 9 . Mycoparasitic Trichoderma species secrete chitinases and glucanases that attack cell wall polymer on other fungi and have been exploited in the development of biocontrol strategies 1 . In this study we describe the .