Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli

CORC > 青岛生物能源与过程研究所 > 中国科学院青岛生物能源与过程研究所 > 材料生物技术研究中心

	Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli
	Cao, Yujin; Xian, Mo; Yang, Jianming; Xu, Xin; Liu, Wei; Li, Liangzhi
刊名	PROTEIN EXPRESSION AND PURIFICATION
	2010-02-01
卷号	69 期号:2 页码:209-214
关键词	GC-MS Heterologous expression ssi2/fab2 Stearoyl-ACP desaturase
ISSN号	ISSN:1046-5928
中文摘要	Fatty acid desaturases are enzymes that introduce double bonds into fatty acyl chains, among which stearoyl-acyl carrier protein desaturase (S-ACP-DES) was widely distributed in the plant kingdom. We cloned the cDNA coding for fab2/ssi2, an S-ACP-DES from Arabidopsis thaliana, into the vector pET30a and heterologously expressed this fatty acid desaturase in Escherichia coli BL21 (M). After being induced with IPTG, the fusion protein was efficiently expressed in a soluble form. The SSI2 desaturase was purified by nickel ion affinity chromatography and the product obtained showed a single band by SDS-PAGE analysis. The expression of ssi2 modified the fatty acid composition of the recombinant strain. The ratio of palmitic acid (16:0) decreased from 45.2% (the control strain) to 35.2% while palmitoleate (16:1 Delta 9) and cis-vaccenate (18:1 Delta 11) levels were enhanced to some extent. The desaturase enzymatic activity was measured in vivo when the enzyme substrate stearic acid was provided in the culture medium. A new fatty acid, oleic acid (18:1 Delta 9)was found in the recombinant strain which did not exist in wild-type E. coli. These results demonstrated that the cofactors of the host system can complement the requirement of the SSI2 desaturase.
英文摘要	Fatty acid desaturases are enzymes that introduce double bonds into fatty acyl chains, among which stearoyl-acyl carrier protein desaturase (S-ACP-DES) was widely distributed in the plant kingdom. We cloned the cDNA coding for fab2/ssi2, an S-ACP-DES from Arabidopsis thaliana, into the vector pET30a and heterologously expressed this fatty acid desaturase in Escherichia coli BL21 (M). After being induced with IPTG, the fusion protein was efficiently expressed in a soluble form. The SSI2 desaturase was purified by nickel ion affinity chromatography and the product obtained showed a single band by SDS-PAGE analysis. The expression of ssi2 modified the fatty acid composition of the recombinant strain. The ratio of palmitic acid (16:0) decreased from 45.2% (the control strain) to 35.2% while palmitoleate (16:1 Delta 9) and cis-vaccenate (18:1 Delta 11) levels were enhanced to some extent. The desaturase enzymatic activity was measured in vivo when the enzyme substrate stearic acid was provided in the culture medium. A new fatty acid, oleic acid (18:1 Delta 9)was found in the recombinant strain which did not exist in wild-type E. coli. These results demonstrated that the cofactors of the host system can complement the requirement of the SSI2 desaturase. (C) 2009 Elsevier Inc. All rights reserved.
学科主题	生物基化学品
WOS标题词	Science & Technology ; Life Sciences & Biomedicine
类目[WOS]	Biochemical Research Methods ; Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
研究领域[WOS]	Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
关键词[WOS]	FATTY-ACID DESATURASE ; MEMBRANE-LIPIDS ; COA DESATURASE ; GENE ; IDENTIFICATION ; CLONING ; PLANTS ; BIOSYNTHESIS ; PURIFICATION ; MUTANTS
收录类别	SCI
语种	英语
WOS记录号	WOS:000272653300013
公开日期	2011-09-09
内容类型	期刊论文
源URL	[http://ir.qibebt.ac.cn//handle/337004/450]
专题	青岛生物能源与过程研究所_材料生物技术研究中心
作者单位	Chinese Acad Sci, Qingdao Inst BioEnergy & Bioproc Technol, Qingdao 266101, Peoples R China
推荐引用方式 GB/T 7714	Cao, Yujin,Xian, Mo,Yang, Jianming,et al. Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli[J]. PROTEIN EXPRESSION AND PURIFICATION,2010,69(2):209-214.
APA	Cao, Yujin,Xian, Mo,Yang, Jianming,Xu, Xin,Liu, Wei,&Li, Liangzhi.(2010).Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli.PROTEIN EXPRESSION AND PURIFICATION,69(2),209-214.
MLA	Cao, Yujin,et al."Heterologous expression of stearoyl-acyl carrier protein desaturase (S-ACP-DES) from Arabidopsis thaliana in Escherichia coli".PROTEIN EXPRESSION AND PURIFICATION 69.2(2010):209-214.