Isolation and partial characterization of novel genes encoding acidic cellulases from metagenomes of buffalo rumens

• Published in: Journal of applied microbiology 2009-07, Vol.107 (1), p.245-256
• Main Authors: Duan, C.-J, Xian, L, Zhao, G.-C, Feng, Y, Pang, H, Bai, X.-L, Tang, J.-L, Ma, Q.-S, Feng, J.-X
• Format: Article
• Language: English
• Subjects: Animals; Bacteria - enzymology; Bacteria - genetics; Biological and medical sciences; Buffaloes; cellulase; cellulases; Cellulases - genetics; Cloning, Molecular - methods; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Fundamental and applied biological sciences. Psychology; Genomic Library; glycosyl hydrolase; metagenome; Metagenomics; Microbiology; properties; Rumen - enzymology; Rumen - microbiology; rumen microorganisms; ruminal microorganisms; Sequence Analysis, DNA
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1111/j.1365-2672.2009.04202.x

To clone and characterize genes encoding novel cellulases from metagenomes of buffalo rumens. A ruminal metagenomic library was constructed and functionally screened for cellulase activities and 61 independent clones expressing cellulase activities were isolated. Subcloning and sequencing of 13 positive clones expressing endoglucanase and MUCase activities identified 14 cellulase genes. Two clones carried two gene clusters that may be involved in the degradation of polysaccharide nutrients. Thirteen recombinant cellulases were partially characterized. They showed diverse optimal pH from 4 to 7. Seven cellulases were most active under acidic conditions with optimal pH of 5·5 or lower. Furthermore, one novel cellulase gene, C67-1, was overexpressed in Escherichia coli, and the purified recombinant enzyme showed optimal activity at pH 4·5 and stability in a broad pH range from pH 3·5 to 10·5. Its enzyme activity was stimulated by dl-dithiothreitol. The cellulases cloned in this work may play important roles in the degradation of celluloses in the variable and low pH environment in buffalo rumen. This study provided evidence for the diversity and function of cellulases in the rumen. The cloned cellulases may at one point of time offer potential industrial applications.