Recombinant expression and purification of human androgen receptor in a baculovirus system.
Recombinant expression and purification of human androgen receptor in a baculovirus system.
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杆状病毒系统中人雄激素受体的重组表达和纯化。
DOI：
10.1006/bbrc.2001.5029
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Dalton,JT
中科院分区：
文献类型：
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作者：
Zhu,Z;Bulgakov,OV;Scott,SS;Dalton,JT
A full-length human androgen receptor (hAR) cDNA was used to produce recombinant baculovirus. Spodoptera frugiperda (Sf9) cells infected with this virus expressed protein with an N-terminal hexahistidine tag (His6-hAR) in soluble and insoluble forms. The soluble cytosolic His6-hAR demonstrated similar association and dissociation half-times for mibolerone, similar binding affinity for mibolerone, and similar steroid specificity as bona fide AR. Under native conditions, the soluble cytosolic His6-hAR was purified to apparent homogeneity in the presence of dihydrotestosterone, using metal ion affinity chromatography. The insoluble pellet fraction was solubilized with strong denaturant 6 M guanidine HCl, and His6-hAR was purified from it in the presence of 6 M guanidine HCl. Both the solubilized crude pellet fraction and the solubilized/purified His6-hAR could be renatured to bind mibolerone. The baculovirus system will therefore provide an efficient means for producing hAR for ligand-binding assays, as well as purifying hAR for detailed molecular analyses.
