Nucleic Acids Research, 2002, Vol. 30, No. 16 e88
© 2002 Oxford University Press
Removal of impurities from transcription factor preparations that alter their DNA-binding properties
Department of Internal Medicine and Department of Molecular Biology, University of Texas–Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8573, USA
*To whom correspondence should be addressed. Tel: +1 214 648 1239; Fax: +1 214 648 1450; Email: thomas.kodadek{at}utsouthwestern.edu
Biochemical studies of transcriptional activators are important for understanding their detailed mechanism of action. Such experiments generally employ chimeric constructs comprised of fused DNA- binding and activation domains that are expressed in, and purified from, Escherichia coli, since full-length activators are usually difficult to express. We report here that such preparations contain chaperone impurities that affect the DNA-binding properties of the activator, for example sharply reducing the half-life of the protein–DNA complex. A simple method to remove these troublesome contaminants is described.