一种用于催产素启动子分析的、快速一步法纯化杆状病毒表达的人雌激素受体的方法。

A rapid one-step method to purify baculovirus-expressed human estrogen receptor to be used in the analysis of the oxytocin promoter.

作者信息

Beekman J M, Cooney A J, Elliston J F, Tsai S Y, Tsai M J

机构信息

Department of Cell Biology, Baylor College of Medicine, Houston, TX 77030.

出版信息

Gene. 1994 Sep 2;146(2):285-9. doi: 10.1016/0378-1119(94)90307-7.

DOI:10.1016/0378-1119(94)90307-7

PMID:8076833

Abstract

We have produced a truncated form of the human estrogen receptor (hER) as a fusion protein with glutathione S-transferase (GST) in Spodoptera frugiperda (Sf) cells using the baculovirus expression vector (BEV) system. The protein is correctly produced and can be purified from crude whole-cell extracts by a single-step, batch-wise affinity-purification procedure. We show that this GST-hER fusion protein binds at its DNA-binding site specifically and in a hormone-inducible manner. Furthermore, we used the purified hER to analyze the complex estrogen response element (ERE) in the promoter of the oxytocin-encoding gene.

摘要

我们利用杆状病毒表达载体（BEV）系统，在草地贪夜蛾（Sf）细胞中制备了一种截短形式的人雌激素受体（hER），它与谷胱甘肽S-转移酶（GST）融合形成融合蛋白。该蛋白能够正确表达，并且可以通过一步批量亲和纯化程序从粗制全细胞提取物中纯化出来。我们发现这种GST-hER融合蛋白能够以激素诱导的方式特异性地结合其DNA结合位点。此外，我们使用纯化的hER来分析催产素编码基因启动子中的复合雌激素反应元件（ERE）。