Screening and Identification of Chitinase Genes from Arctic Marine Bacterium Pseudoalteromonas issachenkonii KOPRI 22718

Abstract

The psychrotolerant Pseudoalteromonas issachenkonii KOPRI 22718 was isolated for high exo-acting chitinase activity in Arctic sea. Previously, an exo-acting chitinase was purified from the culture supernatant and characterized on its biochemical reactions. To identify exochitinase encoding genes, genomic and small-insert DNA library were constructed by using pCC1FOS fosmid/E. coli EPI300 and high-copy plasmid pUC19/ E. coli Rosetta systems, respectively. The transformants from each library were screened on LB/colloidal chitin (0.55%), and finally one superior pUC19 clone was selected for the ability to form clear-zone around the cells. A cluster of three closely linked chitinase genes were revealed as organized in the order of chi III, chi II and chi I on the ~7.5 kb insert DNA. Based on functional analysis for conserved domains, it was predicted that chi III and chi II possess only one catalytic domain and one chitin binding domain, respectively, while chi I has one catalytic domain and one chitin binding domain. Now, chi III is being studied on its over-expression in E. coli and exo-chitinolytic function. This work was supported by a grant to KOPRI (PE11060).