Initiation of in vitro mRNA synthesis from the wild-type lac promoter.

An in vitro transcription system, dependent on catabolite gene activator protein (CAP), utilizing a 200 base-pair restriction fragment, has been used to show that the initiation site of the wild-type Escherichia coli lac mRNA, and that of two mutants, 8d and ps, are identical to that previously reported for the CAP-independent promoter mutant UV5. Order of addition experiments are used to show that the binding of lac repressor to the operator is competitive with that of the RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) to the promoter, thus demonstrating functional overlap of the operator and promoter sites.