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A fast and simple method to prepare the FKBP-rapamycin binding domain of human target of rapamycin for NMR binding assays.

Abstract：

:Mammalian target of rapamycin (TOR) controls cell growth and metabolism in response to the availability of nutrients, growth factors, and the cellular energy status. Misregulation of TOR can result in a pathogenic increase or decrease in organ size and in cancer. TOR can be inhibited by binding of a complex of rapamycin and FKBP to the FKBP-rapamycin binding (FRB) domain. Rapamycin and derivatives of it have been used as immunosuppressive drugs. Because TOR is further an interesting drug target in cancer research, we established an expression, purification, and refolding protocol for the FRB domain of human TOR (hFRB). hFRB is extracted from inclusion bodies, purified by reversed phase HPLC, and refolded by drop-wise dilution of the denatured protein into a native buffer. The procedure is very simple and can easily be scaled up to prepare large amounts of functional protein for high-throughput cancer drug screening assays by NMR and other techniques.

journal_name

Protein Expr Purif

journal_title

Protein expression and purification

authors

Dames SA

doi

10.1016/j.pep.2008.01.014

subject

Has Abstract

pub_date

2008-05-01 00:00:00

pages

31-7

issue

1

eissn

1046-5928

issn

1096-0279

pii

S1046-5928(08)00003-X

journal_volume

59

pub_type

杂志文章

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