Cloning, purification and characterization of a novel GH46 family chitosanase, Csn-CAP, from Staphylococcus capitis. (December 2018)

• Record Type: Journal Article
• Authors: Sun, Huihui; Cao, Rong; Li, Laihao; Zhao, Ling; Liu, Qi
• Is Part Of: Process biochemistry. Volume 75(2018)
• Page Start: 146
• Page End: 151
• Publication Date: 2018-12
• Subjects: Chitosanase -- Chitosan -- Chitooligosaccharide -- Staphylococcus capitis -- Cold-adapted; Biochemical engineering -- Periodicals; Biotechnology -- Periodicals; Biochemistry -- periodicals; Biotechnology -- periodicals; Chemical Engineering -- periodicals; Génie biochimique -- Périodiques; Biotechnologie -- Périodiques; Biochemical engineering; Biotechnology; Periodicals; 660.63
• DOI: 10.1016/j.procbio.2018.09.021
• Languages: English
• ISSNs: 1359-5113

Abstract

Graphical abstract: Highlights: A novel cold-adapted chitosanase Csn-CAP from Staphylococcus capitis was identified. Csn-CAP is a novel member of GH46 family. Csn-CAP exhibits an endo-type cleavage pattern. Csn-CAP hydrolyzes chitosan to yield mainly (GlcN)2 and (GlcN)3 . Abstract: A novel chitosanase gene, designated csn-cap, was cloned from Staphylococcus capitis with codon optimization and functionally expressed in Escherichia coli M15. The recombinant enzyme Csn-CAP, which belongs to the GH46 family, consists of 342 amino acids, which includes a 35-amino acid signal peptide. The mature protein was purified to homogeneity using Ni-NTA affinity chromatography, and the molecular mass of the purified enzyme was estimated to be ∼35 kDa by SDS-PAGE. Csn-CAP displayed a maximal activity at 30 °C and pH 7 and a weak alkaline solution could inhibit its activity harshly. The enzyme was cold-adapted and exhibited 86% of its maximal activity at 20 °C. The level of enzymatic activity was enhanced by some bivalent metal cations, such as Zn 2+, Cu 2+ and especially Mn 2+, which could enhance the activity more than twofold at a 5 mM concentration. The enzyme exhibited strict substrate specificity for chitosan solution, colloidal chitosan and powdery chitosan, but it could not hydrolyze colloidal chitin or carboxymethylcellulose. The thin-layer chromatography result showed that Csn-CAP exhibited an endo-type cleavage pattern and hydrolyzed chitosan to yield mainly (GlcN)2 and (GlcN)3 . Thus, the novel characteristics of the chitosanase Csn-CAP make it a potential candidate for oligosaccharide production-based industries.