Analysis of Interaction of Estradiol with Estrogen Receptor by NMR Spectroscopy
Following binding to estradiol, estrogen receptors (ER) α and ERβ recruit a number of interacting proteins and mediate a plethora of functions. The binding of estrogen with the receptors shows changes in the resonance structure and movement of protons. We cloned ERβ and its trans-activation domain (TAD) and ligand-binding domain (LBD), expressed them in prokaryotic expression vectors, purified them, and studied their interaction with estradiol. In this chapter, a detailed method of preparation of recombinant proteins, SDS-PAGE, silver staining, and NMR are described. Such methods are useful to check the biological activity of bacterially expressed proteins and are applicable to basic and applied research.
Key wordsEstrogenreceptors Cloning SDS-polyacrylamide gel electrophoresis Nuclear magnetic resonance spectroscopy Estradiol
Financial support from UGC-CAS and Department of Biotechnology, Government of India, to M.K.T. is highly acknowledged.
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