微生物学通报 MAY 20, 2008, 35(5): 661~665
Microbiology 2008 by Institute of Microbiology, CAS
tongbao@im.ac.cn
柳广飞* 周集体 周 觅 李敬美
(大连理工大学环境与生命学院 大连 116023)
摘 要: 利用同源建模构建了Rhodobacter sphaeroides的偶氮还原酶AZR的三级结构模型, AZR为一种 型结构的黄素氧化还原蛋白。两类依赖黄素的偶氮还原酶的三级结构对比表明它们具有高度的相似性。在序列和结构对齐分析的基础上, 选择保守位点K109进行K109A和K109H的定点突变研究。突变后K109H的最适pH=6, 而K109A的最适pH=9。突变未改变AZR的最适温度(30℃)。第109位正电荷残基对甲基红的结合有重要影响; 而K109H对NADPH的结合并非保守突变。K109可能只参与对NADPH的2'-磷酸基团的结合, 而对NADH的结合无影响。
关键词: 偶氮还原酶, 同源建模, 定点突变
Structure Modeling of Azoreductase AZR and Site-directed Mutagenesis of Its K109 Residue
LIU Guang-Fei* ZHOU Ji-Ti ZHOU Mi LI Jing-Mei
(Institute of Environmental and Biological Science and Technology, Dalian University of Technology, Dalian 116023)
Abstract: Three-dimensional structure model of azoreductase AZR of Rhodobacter sphaeroides was con-structed using homology modeling method. It is a flavodoxin adopting / structure. Structure alignment of two different types of flavin-dependent azoreductases revealed that they possessed high similarity. Based on sequence and structure analysis, site-directed mutagenesis of K109H and K109A were performed. The opti-mal pH values are pH 6 and pH 9 for K109H and K109A mutant protein, respectively. The optimal tempera-ture (30℃) is not affected by mutagenesis. Positively charged residues at position 109 is necessary for the binding of methyl red, while K109H is not a conserved mutagenesis for the binding of NADPH. K109 may only be involved in the binding of the 2'-phosphate group of NADPH and have no effect on the binding of NADH.
Keywords: Azoreductase, Homology modeling, Site-directed mutagenesis
全文下载：偶氮还原酶AZR的结构及其K109的定点突变研究
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