Expression of active hormone and DNA-binding domains of the chicken progesterone receptor in E. coli - PubMed (original) (raw)

Expression of active hormone and DNA-binding domains of the chicken progesterone receptor in E. coli

J Eul et al. EMBO J. 1989 Jan.

Abstract

Bacterially-expressed fusion proteins containing the DNA-(region C) or hormone-binding (region E) domains of the chicken progesterone receptor (cPR) fused to the C terminus of Escherichia coli beta-galactosidase were analysed for the specificity of interaction with natural and synthetic hormone-responsive elements (HREs) and progestins, respectively. The purified fusion protein containing the progestin-binding domain bound progesterone with an apparent Kd of 1.0-1.5 nM and was specifically photocross-linked with the synthetic progestin R5020 in crude bacterial lysates. Labelling of intact bacterial cells with [3H]R5020 revealed that the majority, if not all, of the bacterially produced hormone-binding domain was active. No differences in the binding to a synthetic palindromic glucocorticoid/progestin-responsive element (GRE/PRE) were found when the bacterially produced cPR DNA-binding domain was compared in methylation interference assays with the full-length chicken progesterone receptor form A expressed in eukaryotic cells. The study of dissociation kinetics, however, revealed differences in the half-life of the complexes formed between the palindromic GRE/PRE and either the receptor form A or the fusion protein containing the cPR DNA-binding domain. DNase I protection experiments demonstrated that the bacterially produced region C of the cPR generated specific 'footprints' on the mouse mammary tumour virus long terminal repeat (MMTV-LTR) which were nearly identical to those previously reported for the rat glucocorticoid receptor.

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References

    1. EMBO J. 1987 Feb;6(2):363-8 - PubMed
    1. Mol Endocrinol. 1987 Nov;1(11):791-801 - PubMed
    1. EMBO J. 1987 May;6(5):1309-15 - PubMed
    1. Nature. 1987 Oct 22-28;329(6141):734-6 - PubMed
    1. Nature. 1987 Dec 3-9;330(6147):444-50 - PubMed

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