Enzymatic properties of a novel chitinase from Chaetomium cupreum

摘要

A gene encoding a novel chitinase chi58 was cloned from the fungus Chaetomium cupreum by using inverse PCR. The DNA sequence of chi58 contains a 1602-bp open reading frame and two introns that are 52 and 201 bp in length. Regarding our silico analysis chi58 is a modular enzyme composed of a family-18 catalytic domain, which is responsible for chitinase activity, and a chitin-binding domain containing several cysteines. Apparently, the function of these domains is to tightly anchor the enzyme onto the large insoluble polymeric substrate. Chi58 has a pI of 4.47 and a deduced molecular mass of 58 kDa. The optimal pH and temperature conditions were determined to be 5.8 and 45℃, respectively, when colloidal chitin was used as the substrate. SDSPAGE and zymogram analyses indicated the presence of a single active chitinase. Cells with pPIC9K-chi58 produced an extracellular chitinase that had an activity of 39 U/ml protein. Metal ions such as Ba2+, Mg2+, K+, Cu2+, Fe3+, Zn2+, and Co2+ also influenced the activity of the recombinant enzyme.