Role of exposed aromatic residues in substrate-binding of CBM family 5 chitin-binding domain of alkaline chitinase

Abstract

Chitinase J (ChiJ) from alkaliphilic Bacillus sp. strain J813 has a multidomain structure containing a catalytic domain (CatD), a fibronectin type III like domain (FnIIID) and a chitin-binding domain (ChBD). It has been shown that the ChBD binds to an insoluble chitin and enhances its degradation by the CatD. Further binding study of the ChBD was performed with a glutathione-S-transferase fusion protein. This fusion protein showed binding abilities to insoluble chitin and chitosan. Two surface-exposed aromatic residues (Trp541 and Trp542) were found in the tertiary-structure model of ChBD and targeted for mutational analysis. Single and double mutations of the two aromatic residues decreased the chitin- and chitosan-binding abilities. It was revealed that these residues would be important for substrate-binding of the ChBD.