CAP and RNA polymerase interactions with the lac promoter: binding stoichiometry and long range effects.
The binding stoichiometries of the complexes formed when the E. coli cyclic AMP receptor protein (CAP) binds to 203 bp lac promoter-operator restriction fragments have been determined. Under quantitative binding conditions, a single dimer of CAP occupies each of two sites in the promoter. Different electrophoretic mobilities are observed for 1:1 complexes formed with L8-UV5 mutant, L305 mutant, and wild type promoter fragments, indicating sequence-specific structural differences between the complexes. The differences in gel mobility between L8-UV5 and wild type complexes disappear when the promoter fragments are cleaved with Hpa II restriction endonuclease. Models in which CAP alters DNA conformation or in which CAP forms a transient intramolecular bridge between two domains of a DNA molecule could account for these observations. The selective binding of RNA polymerase to CAP-promoter complexes is demonstrated: the binding of a single CAP dimer to the promoter is sufficient to stimulate subsequent polymerase binding. Functional CAP molecules are not released from the promoter on polymerase binding.
D002352:Carrier Proteins / Q000235:genetics*; D002373:Cyclic AMP Receptor Protein; D004262:DNA Restriction Enzymes; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D007763:Lac Operon; D009154:Mutation; D011485:Protein Binding; D011953:Receptors, Cyclic AMP / Q000235:genetics*
D002352:Carrier Proteins; D002373:Cyclic AMP Receptor Protein; D011953:Receptors, Cyclic AMP; D012321:DNA-Directed RNA Polymerases; D004262:DNA Restriction Enzymes