DNA binding of the transcription activator protein MeIR from Escherichia coli and its C-terminal domain

MeIR is an Escherichia coli transcription factor belonging to the AraC family. It activates expression of the melAB operon in response to melibiose. Full-length MeIR (MeIR303) binds to two pairs of sites upstream of the melAB transcription start site, denoted sites 1' and 1 and sites 2 and 2', and to a fifth site, R, which overlaps the divergent meIR promoter. The C-terminal domain of MeIR (MeIR173) does not activate transcription. Here we show that, like MeIR303, when MeIR173 binds to sites 1 and 2 It recruits CRP to bind between these sites. Hence, the C-terminal domain is involved in heterologous Interactions. MeIR173 binds to the R site, which has 11 of 18 bp identical to sites 1 and 2 but, surprisingly, does not bind to site 1', which has 12 of 18 bp identical, nor to site 2'. Using electrophoretic mobility shift assays, we show that the binding of MeIR303 to sites 1' and 2' is due to cooperative binding with the adjacent site. This homologous cooperativity requires the N-terminal domain of the protein. Activation of the melAB promoter requires MeIR to occupy site 2', which overlaps the -35 hexamer. Hence, both domains of MeIR are required for transcription activation.