BACTERIAL EXPRESSION OF AN ACTIVE CLASS IB CHITINASE FROM CASTANEA-SATIVA COTYLEDONS

Ch3, an endochitinase of 32 kDa present in Castanea sativa cotyledons, showed in vitro antifungal properties when assayed against Trichoderm a viride. The characterization of a cDNA clone corresponding to this p rotein indicated that Ch3 is a class Ib endochitinase that is synthesi zed as a preprotein with a signal sequence preceding the mature polype ptide. 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 ac tivity secreted from cultured bacteria, which should facilitate the id entification of mutants with altered capacity to hydrolyse chitin.