Molecular cloning, characterization, and expression studies of a novel chitinase gene (ech30) from the mycoparasite Trichoderma atroviride strain P1

• Published in: FEMS microbiology letters 2006-03, Vol.256 (2), p.282-289
• Main Authors: Klemsdal, Sonja S., Clarke, Jihong Liu, Hoell, Ingunn A., Eijsink, Vincent G. H., Brurberg, May-Bente
• Format: Article
• Language: English
• Subjects: 5' Untranslated Regions - genetics; Acetylglucosamine - analogs & derivatives; Acetylglucosamine - pharmacology; Amino Acid Sequence; Biological and medical sciences; Botrytis - growth & development; Chitin; Chitin - pharmacology; Chitinase; Chitinases - antagonists & inhibitors; Chitinases - genetics; Chitinases - isolation & purification; Chitinases - metabolism; Cloning; Cloning, Molecular; confrontation assay; Conserved Sequence; DNA, Fungal - chemistry; DNA, Fungal - genetics; E coli; ech30; Enzyme Inhibitors - pharmacology; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Regulation, Fungal; Growth, nutrition, metabolism, transports, enzymes. Molecular biology; Introns - genetics; Microbiology; Molecular Sequence Data; Molecular Weight; Mycology; Phylogeny; Polymerase chain reaction; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Reverse Transcriptase Polymerase Chain Reaction; RNA, Fungal - analysis; RNA, Messenger - analysis; Sequence Alignment; Sequence Analysis, DNA; Trichoderma; Trichoderma - enzymology; Trichoderma - genetics; Trichoderma atroviride; Trisaccharides - pharmacology
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1111/j.1574-6968.2006.00132.x

Abstract We describe the cloning and characterization of a single copy gene from Trichoderma atroviride P1 encoding a novel 30 kDa chitinase, Ech30. Ech30 is a family 18 chitinase showing low sequence similarity to other Trichoderma chitinases. Real-time quantitative RT-PCR studies revealed that expression of the ech30 gene was induced by the presence of Botrytis cinerea in plate confrontation assays, but hardly by chitin in liquid cultures. Studies of Ech30 purified from an Escherichia coli strain overexpressing the ech30 gene devoid of the leader sequence and a predicted intron, showed that the gene encodes an active chitinase, which, as expected for family 18 chitinases, is inhibited by allosamidin.