%0 Journal Article
%T Saturation mutagenesis of three amino acid positions consisting of the active site of an endoglucanase from termiteCoptotermes formosanus<br>台湾家白蚁内切葡聚糖酶活性中心氨基酸的饱和突变
%A Lihua Lin
%A Guomei Qin
%A Yutuo Wei
%A Liqin Du
%A Zongwen Pang
%A Ribo Huang
%A <br>林丽华
%A 秦国梅
%A 韦宇拓
%A 杜丽琴
%A 庞宗文
%A 黄日波
%J 生物工程学报
%D 2009
%I 
%X Functional improvement to one component of the cellulase, endo-b-1, 4-glucanase, has been a focus of the recent research in this area. We report here the saturation mutagenesis of the active site of an endoglucanase (CfEG) from termite Coptotermes formosanus. First, three dimensional structure of CfEG was built via homology modeling by using a close-related (79% homology in sequence) endo-b-1,4-glucanase (NtEG, PDB id=1ks8) from higher termite Nasutitermes takasagoensis as a template. Second, we identified three corresponding amino acid positions at the active site of CfEG by structural superposition onto NtEG. These three putative amino acids for the active site of CfEG, i.e., Asp53, Asp56 and Glu411, were subjected to saturation mutagenesis using degenerate primers. Among the mutants, Asp53Glu and Asp56Cys showed somewhow higher activities than the wildtype, with the latter having more than 3-fold decrease in Km. Double mutation Asp53Leu/Asp56Ile showed nearly 2-fold increase in specific activity and in the same time 2-fold decrease in Km. Saturation mutagenesis to the position Glu411 produced no active mutant, even changing Glu411 explicitly into its similar amino acids such as Glu411Asp and Glu411Gln could not result in any active mutant. These imply that position Glu411 could be extremely important and therefore indispensable for CfEG functionality.
%K Coptotermes formosanus
%K endoglucanase
%K site-saturation mutagenesis
%K combined mutation
%K site-directed mutagenesis<br>台湾家白蚁
%K 内切葡聚糖酶
%K 定点饱和突变
%K 双突变
%K 定点突变
%U http://www.alljournals.cn/get_abstract_url.aspx?pcid=90BA3D13E7F3BC869AC96FB3DA594E3FE34FBF7B8BC0E591&jid=A66E90C274451689E69F6F0291467824&aid=0ACA205F7D8D490D6BF305F3B4B335F1&yid=DE12191FBD62783C&vid=C5154311167311FE&iid=B31275AF3241DB2D&sid=8B6586F75D2B256A&eid=90EAFEE49150CFCD&journal_id=1000-3061&journal_name=生物工程学报&referenced_num=0&reference_num=10