Bacterial expression of an active class Ib chitinase from Castanea sativa cotyledons |
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Authors: | Isabel Allona Carmen Collada Rosa Casado Javier Paz-Ares Cipriano Aragoncillo |
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Affiliation: | (1) Departamento de Biotecnología. E.T.S. Ingenieros de Montes, Ciudad Universitaria, 28040 Madrid, Spain;(2) Centro Nacional de Biotecnología-CSIC, Campus de Cantoblanco, 28049 Madrid, Spain |
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Abstract: | Ch3, an endochitinase of 32 kDa present in Castanea sativa cotyledons, showed in vitro antifungal properties when assayed against Trichoderma viride. The characterization of a cDNA clone corresponding to this protein indicated that Ch3 is a class Ib endochitinase that is synthesized as a preprotein with a signal sequence preceding the mature polypeptide. Bacterial expression of mature Ch3 fused to the leader peptide of the periplasmic protein ompT resulted in active Ch3 enzyme. A plate assay was adapted for semi-quantitative determination of chitinase activity secreted from cultured bacteria, which should facilitate the identification of mutants with altered capacity to hydrolyse chitin. |
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Keywords: | antifungal protein Castanea sativa chitinase seed defence protein |
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