TRANSCRIPTION ACTIVATION AT CLASS-I FNR-DEPENDENT PROMOTERS - IDENTIFICATION OF THE ACTIVATING SURFACE OF FNR AND THE CORRESPONDING CONTACTSITE IN THE C-TERMINAL DOMAIN OF THE RNA-POLYMERASE ALPHA-SUBUNIT

A library of random mutations in the Escherichia coli fnr gene has bee n screened to identify positive control mutants of FNR that are defect ive in transcription activation at Class I promoters. Single amino aci d substitutions at D43, R72, S73, T118, M120, F181, F186, S187 and F19 1 identify a surface of FNR that is essential for activation which, pr esumably, makes contact with the C-terminal domain of the RNA polymera se or subunit. This surface is larger than the corresponding activatin g surface of the related transcription activator, CRP. To identify the contact surface in the C-terminal domain of the RNA polymerase alpha subunit, a library of mutations in the rpoA gene was screened for or m utants that interfered with transcription activation at Class I FNR-de pendent promoters. Activation was reduced by deletions of the alpha C- terminal domain, by substitutions known to affect DNA binding by alpha , by substitutions at E261 and by substitutions at L300, E302, D305, A 308, G315 and R317 that appear to identify contact surfaces of alpha t hat are likely to make contact with FNR at Class I promoters. Again, t his surface differs from the surface used by CRP at Class I CRP-depend ent promoters.