A STREPTOMYCES CHITOSANASE IS ACTIVE IN TRANSGENIC TOBACCO

Growth inhibition towards Rhizopus nigricans, Fusarium oxysporum f. sp . radicis-lycopersici, Verticillium albo-atrum and Pythium ultimum was observed in vitro using a purified chitosanase from an actinomycete, Streptomyces sp. strain N174. The corresponding gene, with its own sig nal peptide, was inserted into pBI121.7 shuttle vector to transform to bacco. Transgenic plants were analysed for chitosanase activity by a s odium dodecyl sulfate-polyacrylamide gel electrophoresis assay. Two ma jor and one minor active electrophoretic forms were detected in transg enic tobacco. Some chitosanases were recovered not only in leaf homoge nates but also in leaf intercellular fluid extracts. One chitosanase e lectrophoretic form migrated very closely to the purified Streptomyces mature protein while die others corresponded to molecules of higher m olecular mass. The N-terminus sequence was determined for one of the t hree chitosanase forms. It exhibited a different signal peptide cleava ge site when compared to the mature chitosanase from Streptomyces. Thi s is the first report oil the expression of an active chitosanase gene with antimicrobial potential in plants.