Over-expression and purification of isotopically labeled recombinant ligand-binding domain of orphan nuclear receptor human B1-binding factor/human liver receptor homologue 1 for NMR studies

	Over-expression and purification of isotopically labeled recombinant ligand-binding domain of orphan nuclear receptor human B1-binding factor/human liver receptor homologue 1 for NMR studies
	Chen, X; Tong, XT; Xie, YH; Wang, Y; Ma, JB; Gao, DM; Wu, HM; Chen, HB
刊名	PROTEIN EXPRESSION AND PURIFICATION
	2006
卷号	45 期号:1 页码:99-106
关键词	hB1F/hLRH-1 isotope labeling M9 minimal medium NMR NR5A2 protein over-expression purification recombinant LBD
通讯作者	Wu, HM (reprint author), Chinese Acad Sci, Shanghai Inst Organ Chem, State Key Lab Bioorgan & Nat Prod Chem, 345 Lingling Lu, Shanghai 200032, Peoples R China.,hmwu@mail.sioc.ac.cn
英文摘要	The human hepatitis B virus enhancer II B1 binding factor (hB1F), which regulates the expression of hepatitis B virus genes, is identified as a nuclear receptor. It regulates several liver-specific genes and plays an important role in the bile acid biosynthesis pathway. A significantly optimized protocol has been worked out to prepare N-15 and/or C-13-labeled hB1F ligand-binding domain in minimal medium with high yields for NMR studies. Under the various conditions optimized for the purification of His(6)-hB1F ligand-binding domain, the yield of the purified protein is estimated to be 25-30 mg from 0.5 L of M9 minimal media. Electrospray ionization mass spectrometry data confirm the correctness of the primary sequence. Dynamic light scattering experiment proves that the protein exists as a monomeric form. In addition, the circular dichroism results show that the protein has a well-regulated secondary structure and a high alpha-helical content in ammonium bicarbonate buffer at 20 degrees C and pH 7.4. Finally, uniformly N-15-labeled protein is characterized by a TROSY-HSQC spectrum, and the dispersion of N-15-H-1 cross-peaks in the spectrum indicates the presence of well-ordered and properly folded protein as a monomer. (C) 2005 Elsevier Inc. All rights reserved.
学科主题	Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
类目[WOS]	Biochemical Research Methods ; Biochemistry & Molecular Biology ; Biotechnology & Applied Microbiology
关键词[WOS]	FETOPROTEIN TRANSCRIPTION FACTOR ; HOMEODOMAIN PROTEIN FTZ ; HEPATITIS-B-VIRUS ; DIFFERENTIAL EXPRESSION ; AROMATASE EXPRESSION ; LRH-1 ; GENE ; SUPERFAMILY ; SENSITIVITY ; ACTIVATION
收录类别	SCI
语种	英语
WOS记录号	WOS:000234521100013
内容类型	期刊论文
版本	出版稿
源URL	[http://202.127.25.143/handle/331003/1661]
专题	上海生化细胞研究所_上海生科院生化细胞研究所
推荐引用方式 GB/T 7714	Chen, X,Tong, XT,Xie, YH,et al. Over-expression and purification of isotopically labeled recombinant ligand-binding domain of orphan nuclear receptor human B1-binding factor/human liver receptor homologue 1 for NMR studies[J]. PROTEIN EXPRESSION AND PURIFICATION,2006,45(1):99-106.
APA	Chen, X.,Tong, XT.,Xie, YH.,Wang, Y.,Ma, JB.,...&Chen, HB.(2006).Over-expression and purification of isotopically labeled recombinant ligand-binding domain of orphan nuclear receptor human B1-binding factor/human liver receptor homologue 1 for NMR studies.PROTEIN EXPRESSION AND PURIFICATION,45(1),99-106.
MLA	Chen, X,et al."Over-expression and purification of isotopically labeled recombinant ligand-binding domain of orphan nuclear receptor human B1-binding factor/human liver receptor homologue 1 for NMR studies".PROTEIN EXPRESSION AND PURIFICATION 45.1(2006):99-106.