Study on a Specific Site Tyr^(444) on a Hyperthermophilic Enzyme APE1547  

作　　者：RAO Lang BI Yun-feng XIE Gui-qiu ZHANG Fei WANG Yan CAO Shu-gui GAO Ren-jun 

机构地区：[1]Key Laboratory for Molecular Enzymology and Engineering, Ministry of Education, Jilin University, Changchun 130023, P R. China [2]College of Food Science and Engineering, Jilin Agricultural University, Changchun 130118, P. R. China [3]School of Pharmacy, Jilin University, Changchun 130021, P. R. China

出　　处：《Chemical Research in Chinese Universities》2009年第3期353-356,共4页高等学校化学研究（英文版）

基　　金：Supported by the National Natural Science Foundation of China(Nos.30400081 and 20432010)

摘　　要：Hyperthermophilic enzyme APE1547 is an extremely thermostable recombinant protein from thermophilic archaeon Aeropyrumpernix K1. The Tyr444 located in the catalytic domain adjacent to the catalytic amino acid Ser445 and formed hydrogen bond with Ile567. To study the effect of Tyr444 on the activity of APE1547, site-directed mutagenesis was applied. Two mutant enzymes T444S and T444G were created. Comparison of the mutant enzymes with wide enzyme, the thermostability of mutants T444S and T444G decreased by 10%-20%, but the catalytic efficiency of mutants toward pNPC8 and Ac-Leu-pNA increased 1.33 and 1.75 fold respectively. Molecular modeling shows that the elimination of hydrogen bond between Tyr444 and Ile567 is the cause of the decrease in thermostability and increase in catalytic efficiency. These observations suggest that Tyr444 plays an important role in the catalytic ability and thermostability of this enzyme.Hyperthermophilic enzyme APE1547 is an extremely thermostable recombinant protein from thermophilic archaeon Aeropyrumpernix K1. The Tyr444 located in the catalytic domain adjacent to the catalytic amino acid Ser445 and formed hydrogen bond with Ile567. To study the effect of Tyr444 on the activity of APE1547, site-directed mutagenesis was applied. Two mutant enzymes T444S and T444G were created. Comparison of the mutant enzymes with wide enzyme, the thermostability of mutants T444S and T444G decreased by 10%-20%, but the catalytic efficiency of mutants toward pNPC8 and Ac-Leu-pNA increased 1.33 and 1.75 fold respectively. Molecular modeling shows that the elimination of hydrogen bond between Tyr444 and Ile567 is the cause of the decrease in thermostability and increase in catalytic efficiency. These observations suggest that Tyr444 plays an important role in the catalytic ability and thermostability of this enzyme.

关 键 词：Hyperthermophilic enzyme APE 1547 Site-directed mutagenesis Thermostability ACTIVITY 

分 类 号：Q517[生物学—生物化学] TQ925[轻工技术与工程—发酵工程]