title: Identification of activity related amino acid mutations of a GH9 termite cellulase.
abstract: Site-directed mutagenesis of the endo-beta-1,4-glucanase (EG) gene from the termite Reticulitermes speratus EG (RsEG) was performed to gain a better understanding of the role of certain amino acid residues in the activity of the enzyme. Three mutants, G91A, Y97W and K429A had higher activities towards carboxymethyl cellulose than the wild type. The mutations had synergistic effects since each single mutant exhibited about 3-4-fold of wild type activity, but the corresponding activities for double and triple mutants were 7-13-fold. Mutant G147R lost the enzymatic activity completely, suggesting G147 plays a significant role in maintaining enzyme activity. The predicted roles of Asp53, Asp56 and Glu411 in enzymatic catalysis were experimentally verified since the resultant mutants lost the enzyme activities. This study presents the first report on the relationship between amino acid residues and enzyme activity of termite EG, and the information will potentially be useful for industrial application of termite-origin cellulase.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong, PR China. jinfgni@sdu.edu.cn
authors.forename: Jinfeng
authors.initials: J
authors.lastname: Ni
authors.forename: Motomi
authors.initials: M
authors.lastname: Takehara
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Watanabe
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000235:genetics*; D000818:Animals; D055162:Biocatalysis; D020134:Catalytic Domain; D002480:Cellulase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D020049:Isoptera; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017386:Sequence Homology, Amino Acid
chemical_list: D000596:Amino Acids; D002480:Cellulase