CYTR CAMP-CRP NUCLEOPROTEIN FORMATION IN ESCHERICHIA-COLI - THE CYTR REPRESSOR BINDS ITS OPERATOR AS A STABLE DIMER IN A TERNARY COMPLEX WITH CAMP-CRP/

The CytR repressor protein relies on protein-protein interactions to t he cAMP-CRP complex to bind its operators with sufficiently high affin ity to repress transcription. Here, the quaternary structure of CytR a nd the mechanism underlying the cooperative binding of CytR and cAMP-C RP have been analyzed. Using a modified Ferguson analysis in which pro tein-DNA complexes are separated in a non-denaturing gel system, we sh ow that CytR binds its operators as a dimer alone as well as in a tern ary complex with cAMP-CRP. Analyses of DNA binding of CytR at low prot ein concentrations indicate that CytR is a dimer in solution at physio logical concentrations. Moreover, the CytR inducer cytidine was found not to have any effect on the oligomerization of free CytR or DNA boun d CytR. Thus, these data rule out the possibility that the cooperative DNA binding of CytR and cAMP-CRP involves induced dimerization of Cyt R, and they suggest that cytidine interrupts the cooperative binding o f CytR and cAMP-CRP solely by perturbing the protein-protein interacti ons between the two proteins. (C) 1996 Academic Press Limited