Functional characterization of a thermostable endoglucanase belonging to glycoside hydrolase family 45 from Fomitopsis palustris

• Published in: Applied microbiology and biotechnology 2018-08, Vol.102 (15), p.6515-6523
• Main Authors: Cha, Ju-Hee, Yoon, Jeong-Jun, Cha, Chang-Jun
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Basidiomycota; Biomedical and Life Sciences; Biotechnologically Relevant Enzymes and Proteins; Biotechnology; Brown rot; Carboxymethyl cellulose; Carboxymethylcellulose; Catalysis; Cellulase - genetics; Cellulase - metabolism; Cellulose; Cellulose - chemistry; Cellulose - metabolism; Cloning, Molecular; Coriolaceae - enzymology; Endoglucanase; Enzymatic activity; Enzymes; Fungi; Glucan; Glucans; Glycoside hydrolase; Hydrolase; Industrial applications; Industrial Microbiology; Life Sciences; Microbial Genetics and Genomics; Microbiology; Mutagenesis, Site-Directed; Phosphoric acid; Physiological aspects; Pichia - genetics; Pichia pastoris; Proteins; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Site-directed mutagenesis; Substrate Specificity; Substrates; Yeast; β-Glucan
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-018-9075-5

A gene encoding an endoglucanase belonging to subfamily C of glycoside hydrolase family 45 (GH45) was identified in the brown rot fungus Fomitopsis palustris and functionally expressed in Pichia pastoris . The recombinant protein displayed hydrolytic activities toward various substrates such as carboxymethyl cellulose, phosphoric acid swollen cellulose, glucomannan, lichenan, and β-glucan. In particular, the enzyme had a unique catalytic efficiency on β-1,4-glucans rather than mixed β-1,3/1,4-glucans as compared to other GH45 endoglucanases. The fungal enzyme was relatively thermostable, retaining more than 91.4% activity at 80 °C for 1 h. Site-directed mutagenesis studies revealed that the mutants N95D and D117N had significantly reduced enzymatic activities, indicating that both residues are essential for the catalytic reaction. Our study expands knowledge and understanding of the catalytic mechanism of GH45 subfamily C enzymes and also suggests that this thermostable endoglucanase from F. palustris has great potential in industrial applications.