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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; purification; recombinant LBD
Alternative Title同位素标记的人核激素受体hB1F配体结合域的表达和纯化
Chen X(陈翔); Tong XT(童孝田); Xie YH(谢幼华); Wang Y(汪垣); Ma JB(麻锦彪); Gao DM(高大明); Wu HM(吴厚铭); Chen HB(陈海宝)
2006-01-01
Source PublicationProtein Expr. Purif.
ISSN1046-5928
Volume45Issue:1Pages:99-106
AbstractThe 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.
Subject Area天然产物有机化学
Department上海有机化学研究所生命有机化学国家重点实验室; 上海生物化学与细胞生物学研究所分子生物学国家重点实验室
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Indexed BySCI
Language英语
Document Type期刊论文
Identifierhttp://ir.sioc.ac.cn/handle/331003/17375
Collection上海有机化学研究所
Corresponding AuthorWu HM(吴厚铭); Chen HB(陈海宝)
Recommended Citation
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 over-expression; purification; recombinant LBD[J]. Protein Expr. Purif.,2006,45(1):99-106.
APA 陈翔.,童孝田.,谢幼华.,汪垣.,麻锦彪.,...&陈海宝.(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 over-expression; purification; recombinant LBD.Protein Expr. Purif.,45(1),99-106.
MLA 陈翔,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 over-expression; purification; recombinant LBD".Protein Expr. Purif. 45.1(2006):99-106.
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