Molecular cloning and characterization of a β-glucanase from Piromyces rhizinflatus
Journal
Journal of Bioscience and Bioengineering
Journal Volume
111
Journal Issue
5
Pages
541-546
Date Issued
2011
Author(s)
Abstract
Cellulose is the most abundant renewable polysaccharide with a high potential for degradation to useful end products. In nature, most cellulose is produced as crystalline cellulose. Therefore, cellulases with high hydrolytic activity against crystalline cellulose are of great interest. In this study, a crystalline cellulose degradation enzyme was investigated. The cDNA encoding a β-glucanase, CbhYW23-2, was cloned from the ruminal fungus Piromyces rhizinflatus. To examine the enzyme activities, CbhYW23-2 was expressed in Escherichia coli as a recombinant His6 fusion protein and purified by immobilized metal ion-affinity chromatography. Response surface modeling (RSM) combined with central composite design (CCD) and regression analysis was then employed for the planned statistical optimization of the β-glucanase activities of CbhYW23-2. The optimal conditions for the highest β-glucanase activity of CbhYW23-2 were observed at 46.4°C and pH 6.0. The results suggested that RSM combined with CCD and regression analysis were effective in determining optimized temperature and pH conditions for the enzyme activity of CbhYW23-2. CbhYW23-2 also showed hydrolytic activities toward Avicel, carboxymethyl cellulose (CMC), lichenan, and pachyman. The results also proved that the high activity of CbhYW23-2 on crystalline cellulose makes it a promising candidate enzyme for biotechnological and industrial applications. © 2011 The Society for Biotechnology, Japan.
Subjects
β-Glucanase; Optimization; Piromyces rhizinflatus; Response surface methodology
SDGs
Other Subjects
Carboxymethyl cellulose; cDNA encoding; Central composite designs; Crystalline cellulose; End-products; Fusion proteins; Glucanase; High activity; High potential; Hydrolytic activities; Immobilized metals; Molecular cloning; Optimal conditions; pH condition; Piromyces rhizinflatus; Response surface methodology; Response surface modeling; Statistical optimization; Affinity chromatography; Cellulose; Cloning; Crystalline materials; Degradation; Enzymes; Escherichia coli; Industrial applications; Metal ions; Optimization; Regression analysis; Surface properties; Enzyme activity; beta glucan hydrolase; carboxymethylcellulose; microcrystalline cellulose; article; biotechnology; enzyme activity; hydrolysis; immobilized metal affinity chromatography; molecular cloning; nonhuman; nucleotide sequence; pH measurement; Piromyces; Piromyces rhizinflatus; temperature sensitivity; Amino Acid Sequence; Cellulases; Cellulose; Cloning, Molecular; Escherichia coli; Fungal Proteins; Glucans; Hydrogen-Ion Concentration; Hydrolysis; Molecular Sequence Data; Piromyces; Recombinant Proteins; Substrate Specificity; Temperature; Escherichia coli; Fungi; Piromyces rhizinflatus
Type
journal article