Functional Analysis of the Chitin-binding Domain of a Family 19 Chitinase from Streptomyces griseus HUT6037: Substrate-binding Affinity and cis-Dominant Increase of Antifungal Function

Abstract

  Chitinase C (ChiC) is the first bacterial family 19 chitinase discovered in Streptomyces griseus HUT6037. While it shares significant similarity with the plant family 19 chitinases in the catalytic domain, its N-terminal chitin-binding domain (ChBD_ChiC) differs from those of the plant enzymes. ChBD_ChiC and the catalytic domain (CatD_ChiC), as well as intact ChiC, were separately produced in E. coli and purified to homogeneity. Binding experiments and isothermal titration calorimetry assays demonstrated that ChBD_ChiC binds to insoluble chitin, soluble chitin, cellulose, and N-acetylchitohexaose (roughly in that order). A deletion of ChBD_ChiC resulted in moderate (about 50%) reduction of the hydrolyzing activity toward insoluble chitin substrates, but most (about 90%) of the antifungal activity against Trichoderma reesei was abolished by this deletion. Thus, this domain appears to contribute more importantly to antifungal properties than to catalytic activities. ChBD_ChiC itself did not have antifungal activity or a synergistic effect on the antifungal activity of CatD_ChiC in trans.