Binding of the cyclic AMP receptor protein of Escherichia coli to RNA polymerase.
Fluorescence polarization studies were used to study the interaction of a fluorescein-labelled conjugate of the Escherichia coli cyclic AMP receptor protein (F-CRP) and RNA polymerase. Under conditions of physiological ionic strength, F-CRP binds to RNA polymerase holoenzyme in a cyclic AMP-dependent manner; the dissociation constant was about 3 microM in the presence of cyclic AMP and about 100 microM in its absence. Binding to core RNA polymerase under the same conditions was weak (Kdiss. approx. 80-100 microM) and independent of cyclic AMP. Competition experiments established that native CRP and F-CRP compete for the same binding site on RNA polymerase holoenzyme and that the native protein binds about 3 times more strongly than does F-CRP. Analytical ultracentrifuge studies showed that CRP binds predominantly to the monomeric rather than the dimeric form of RNA polymerase.
Pinkney
M
M
Department of Biology, University of York, U.K.
Hoggett
J G
JG
D001051:Apoenzymes / Q000378:metabolism; D001667:Binding, Competitive; D003067:Coenzymes / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D046911:Macromolecular Substances; D009994:Osmolar Concentration; D011953:Receptors, Cyclic AMP / Q000378:metabolism*; D013861:Thiocyanates
D001051:Apoenzymes; D003067:Coenzymes; D005452:Fluoresceins; D005456:Fluorescent Dyes; D046911:Macromolecular Substances; D011953:Receptors, Cyclic AMP; D013861:Thiocyanates; D000242:Cyclic AMP; D012321:DNA-Directed RNA Polymerases; D016650:Fluorescein-5-isothiocyanate