Family 19 chitinases from Streptomyces thermoviolaceus OPC-520: Molecular cloning and characterization

Family 19 chitinase genes, chi35 and chi25 of Streptomyces thermoviolaceus OPC-520, were cloned and sequenced. The chi35 and chi25 genes were arranged in tandem and encoded deduced proteins of 39,762 and 28,734 Da, respective ly. Alignment of the deduced amino acid sequences demonstrated that Chi35 h as an N-terminal domain and a catalytic domain and that Chi25 is an enzyme consisting of only a catalytic domain. Amino acid sequences of the catalyti c domains of both enzymes, which are highly similar to each other, suggeste d that these enzymes belong to the family 19 chitinases. The cloned Chi35 a nd Chi25 were purified from E. coli and S. lividans as a host, respectively . The optimum pH of Chi35 and Chi25 were 5-6, and the optimum temperature o f Chi35 and Chi25 were 60 and 70 degreesC, respectively. Chi35 bound to chi tin, Avicel, and xylan. On the other hand, Chi25 bound to these polysacchar ides more weakly than did Chi35. These results indicate that the N-terminal domain of Chi35 functions as a polysaccharide-binding domain. Furthermore, Chi35 showed more efficient hydrolysis of insoluble chitin and stronger an tifungal activity than Chi25, In the polysaccharide-binding domain of Chi35 , there are three reiterated amino acid sequences starting from C-L-D and e nding,vith W, and the repeats were similar to xylanase (STX-I) from the sam e strain. However, the repeats did not show sequence similarity to any of t he known chitin-binding domains and cellulose-binding domains.