Cloning and overexpression of a new chitosanase gene from Penicillium sp. D-1

Abstract

A chitosanase gene, csn, was cloned from Penicillium sp. D-1 by inverse PCR. The cDNA sequence analysis revealed that csn had no intron. The deduced CSN protein consists of 250 amino acids including a 20-amino acid signal peptide, and shared 83.6% identity with the family 75 chitosanase from Talaromyces stipitatus (B8M2R4). The mature protein was overexpressed in Escherichia coli and purified with the affinity chromatography of Ni2+-NTA. The novel recombinant chitosanase showed maximal catalytic activity at pH 4.0 and 48°C. Moreover, the activity of CSN was stable over a broad pH range of 3.0-8.0, and the enzymatic activity was significantly enhanced by Mg2+ and Mn2+. The CSN could effectively hydrolyze colloidal chitosan and chitosan, while could not hydrolyze chitin and carboxymethylcellulose (CMC). Due to the particular acidophily, CSN has the potential application in the recycling of chitosan wastes.The GenBank/EMBL/DDBJ accession numbers for the 18S rRNA gene and chitosanase gene of strain D-1 are JF950269 and JF950270, respectively.

Marine‐polysaccharide degrading enzymes: Status and prospects

“…Most bacterial chitosanases belong to family GH46, and members of this family have been extensively studied in terms of their catalytic mechanisms and protein structures (Saito et al., 1999). Family GH75 includes chitosanases from fungi (Zhu et al., 2012). Chitosanases from families GH46, GH75, and GH80 are believed to be substrate specific (limited to chitosan), while GH5, GH7, and GH8 chitosanases also tend to possess other enzyme activities, such as cellulase and transglycosylation activities (Ike et al., 2007; Pelletier & Sygusch, 1990; Tanabe, Morinaga, Fukamizo, & Mitsutomi, 2003).…”

Structural and biochemical insights into the degradation mechanism of chitosan by chitosanase OU01

“…Chitosanases are found in multiple organisms, including fungi and bacteria, and have been classified into six glycoside hydrolase (GH) families: GH5,7,8,46, 75 and 80 [12][13][14]. The catalytic mechanism and three-dimensional protein structures have been well studied, especially for chitosanases from the GH46 family [10,[14][15][16][17].…”

Molecular characterization of an endo-type chitosanase from the fish pathogenRenibacteriumsp. QD1

“…The activation effect of Mn 2+ was also observed in chitosanase of Penicillium sp. D-1 (Zhu et al ., 2012), but the opposite was observed in Pseudomonas sp. TKU015 (Wang et al , 2008) and Serratia sp .…”