Semi-rational engineering of cytochrome CYP153A from Marinobacter aquaeolei for improved omega-hydroxylation activity towards oleic acid

CORC > 天津工业生物技术研究所 > 生物催化与绿色化工朱敦明 > 期刊论文

	Semi-rational engineering of cytochrome CYP153A from Marinobacter aquaeolei for improved omega-hydroxylation activity towards oleic acid
	Duan, Yan 1; Ba, Lina 1,2; Gao, Jianwei 1; Gao, Xianxing 1; Zhu, Dunming 3; de Jong, Rene M.4; Mink, Daniel 5; Kaluzna, Iwona 5; Lin, Zhanglin 1
刊名	APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
	2016-10-01
卷号	100 期号:20 页码:8779-8788
关键词	Cytochrome CYP153A Oleic acid Electron transfer Site-directed saturation mutagenesis Iterative saturation mutagenesis
英文摘要	omega-Hydroxy oleic acid is an important intermediate for the synthesis of certain polyesters and polyamides. In this study, a functional CYP153A/ putidaredoxin (Pdx)/putidaredoxin reductase (Pdr) hybrid system was engineered for improved omega-hydroxylation activity towards oleic acid. By the combination of site-directed saturation mutagenesis (SDSM) and iterative saturation mutagenesis (ISM), a best mutant (Variant II) was obtained with mutations at two sites (S120 and P165) at the Pdx interaction interface with CYP153A, and one site (S453) in the substrate binding pocket. The in vitro-reconstituted activity of Variant II with purified Pdx and Pdr was 2.7-fold that of the template, while the whole cell transformation activity was 2.0-fold that of the template. A 96-well format-based screening scheme for CYP153A was also developed, which should be useful for engineering of other P450s with low activity. Kinetic analyses indicated that the activity improvement for CYP153A variants largely resulted from enhanced electron transfer. This further demonstrates the importance of the electron transfer between P450s and the non-native redox partners for the overall performance of hybrid P450 systems.
WOS标题词	Science & Technology ; Life Sciences & Biomedicine
类目[WOS]	Biotechnology & Applied Microbiology
研究领域[WOS]	Biotechnology & Applied Microbiology
关键词[WOS]	ENHANCED CATALYTIC-ACTIVITY ; FATTY-ACIDS ; DIRECTED EVOLUTION ; ELECTRON-TRANSFER ; ESCHERICHIA-COLI ; SYNTHETIC APPLICATION ; SELF-SUFFICIENT ; SYSTEM ; MONOOXYGENASES ; EXPRESSION
收录类别	SCI
语种	英语
WOS记录号	WOS:000385128000014
内容类型	期刊论文
源URL	[http://124.16.173.210/handle/834782/2946]
专题	天津工业生物技术研究所_生物催化与绿色化工朱敦明_期刊论文
作者单位	1.Tsinghua Univ, Natl Engn Lab Ind Enzymes, Dept Chem Engn, One Tsinghua Garden Rd, Beijing 100084, Peoples R China 2.Shanghai Acad Agr Sci, Inst Edible Fungi, 1000 Jinqi Rd, Shanghai 201403, Peoples R China 3.Chinese Acad Sci, Natl Engn Lab Ind Enzymes, Tianjin Inst Ind Biotechnol, Tianjin Airport Econ Area, 32 Xi Qi Dao, Tianjin 300308, Peoples R China 4.DSM Biotechnol Ctr, Alexander Fleminglaan 1, NL-2613 AX Delft, Netherlands 5.DSM Ahead R&D BV Innovat Synth, Urmonderbaan 22, NL-6167 RD Geleen, Netherlands
推荐引用方式 GB/T 7714	Duan, Yan,Ba, Lina,Gao, Jianwei,et al. Semi-rational engineering of cytochrome CYP153A from Marinobacter aquaeolei for improved omega-hydroxylation activity towards oleic acid[J]. APPLIED MICROBIOLOGY AND BIOTECHNOLOGY,2016,100(20):8779-8788.
APA	Duan, Yan.,Ba, Lina.,Gao, Jianwei.,Gao, Xianxing.,Zhu, Dunming.,...&Lin, Zhanglin.(2016).Semi-rational engineering of cytochrome CYP153A from Marinobacter aquaeolei for improved omega-hydroxylation activity towards oleic acid.APPLIED MICROBIOLOGY AND BIOTECHNOLOGY,100(20),8779-8788.
MLA	Duan, Yan,et al."Semi-rational engineering of cytochrome CYP153A from Marinobacter aquaeolei for improved omega-hydroxylation activity towards oleic acid".APPLIED MICROBIOLOGY AND BIOTECHNOLOGY 100.20(2016):8779-8788.