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Computation-Aided?Rational?Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 ­Glucuronidase

文献类型：期刊论文

;
作者Han, Beijia; Hou, Yuhui; Jiang, Tian; Lv, Bo; Zhao, Lina; Feng, Xudong; Li, Chun; Zhao LN(赵丽娜)
刊名JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY ; JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
出版日期2018 ; 2018
卷号66期号:43页码:11380-11389
关键词beta-glucuronidase beta-glucuronidase molecular dynamics simulation protein engineering C-terminal region glycyrrhetinic acid molecular dynamics simulation protein engineering C-terminal region glycyrrhetinic acid
ISSN号0021-8561 ; 0021-8561
DOI10.1021/acs.jafc.8b03449 ; 10.1021/acs.jafc.8b03449
文献子类Article ; Article
英文摘要In this study, computation-aided design on the basis of structural analysis was employed to rationally identify a highly dynamic C-terminal region that regulates the stability, expression level, and activity of a GH2 fungal glucuronidase from Aspergillus oryzae Li-3 (PGUS). Then, four mutants with a precisely truncated C-terminal region in different lengths were constructed; among them, mutant D591-604 with a 3.8-fold increase in half-life at 65 degrees C and a 6.8 kJ/mol increase in Gibbs free energy showed obviously improved kinetic and thermodynamic stability in comparison to PGUS. Mutants D590-604 and D591-604 both showed approximately 2.4-fold increases in the catalytic efficiency k(cat)/K-m and 1.8-fold increases in the expression level. Additionally, the expression level of PGUS was doubled through a C-terminal region swap with bacterial GUS from E. coli (EGUS). Finally, the robust PGUS mutants D590-604 and D591-604 were applied in the preparation of glycyrrhetinic acid with 4.0- and 4.4-fold increases in concentration through glycyrrhizin hydrolysis by a fed-batch process.; In this study, computation-aided design on the basis of structural analysis was employed to rationally identify a highly dynamic C-terminal region that regulates the stability, expression level, and activity of a GH2 fungal glucuronidase from Aspergillus oryzae Li-3 (PGUS). Then, four mutants with a precisely truncated C-terminal region in different lengths were constructed; among them, mutant D591-604 with a 3.8-fold increase in half-life at 65 degrees C and a 6.8 kJ/mol increase in Gibbs free energy showed obviously improved kinetic and thermodynamic stability in comparison to PGUS. Mutants D590-604 and D591-604 both showed approximately 2.4-fold increases in the catalytic efficiency k(cat)/K-m and 1.8-fold increases in the expression level. Additionally, the expression level of PGUS was doubled through a C-terminal region swap with bacterial GUS from E. coli (EGUS). Finally, the robust PGUS mutants D590-604 and D591-604 were applied in the preparation of glycyrrhetinic acid with 4.0- and 4.4-fold increases in concentration through glycyrrhizin hydrolysis by a fed-batch process.
电子版国际标准刊号1520-5118 ; 1520-5118
WOS关键词CANCER DRUG TOXICITY ; CANCER DRUG TOXICITY ; IN-SILICO DESIGN ; DIRECTED EVOLUTION ; ENGINEERING HYPERTHERMOSTABILITY ; GLYCYRRHIZIC ACID ; AMINO-ACIDS ; THERMOSTABILITY ; BINDING ; SITE ; XYLANASE ; IN-SILICO DESIGN ; DIRECTED EVOLUTION ; ENGINEERING HYPERTHERMOSTABILITY ; GLYCYRRHIZIC ACID ; AMINO-ACIDS ; THERMOSTABILITY ; BINDING ; SITE ; XYLANASE
WOS研究方向Biophysics ; Biophysics ; Chemistry ; Materials Science ; Chemistry ; Materials Science
语种英语 ; 英语
WOS记录号WOS:000449239400021 ; WOS:000449239400021
源URL[http://ir.ihep.ac.cn/handle/311005/286706]  
专题高能物理研究所_多学科研究中心
作者单位中国科学院高能物理研究所
推荐引用方式
GB/T 7714		 Han, Beijia,Hou, Yuhui,Jiang, Tian,et al. Computation-Aided?Rational?Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 ­Glucuronidase, Computation-Aided?Rational?Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 ­Glucuronidase[J]. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY, JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY,2018, 2018,66, 66(43):11380-11389, 11380-11389.
APA Han, Beijia.,Hou, Yuhui.,Jiang, Tian.,Lv, Bo.,Zhao, Lina.,...&赵丽娜.(2018).Computation-Aided?Rational?Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 ­Glucuronidase.JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY,66(43),11380-11389.
MLA Han, Beijia,et al."Computation-Aided?Rational?Deletion of C-Terminal Region Improved the Stability, Activity, and Expression Level of GH2 ­Glucuronidase".JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 66.43(2018):11380-11389.

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来源：高能物理研究所

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