Domain Organization of the RNA Polymerase σ70Subunit

We used limited trypsin digestion to determine the domain organization of theEscherichia coliRNA polymerase σ70subunit. Trypsin-resistant fragments containing σ70conserved region 2 (σ702), and carboxy-terminal fragments containing conserved regions 3 and 4 (σ703 – 4) were identified by a combination of amino acid sequencing and mass spectrometry. The domains were studied for partial biochemical functions of σ70·σ702bound core RNA polymerase competitively with intact σ70. In contrast to σ702alone, the RNA polymerase holoenzyme formed with σ702specifically bound a single-stranded DNA oligomer with a sequence corresponding to the non-template strand of the −10 promoter element (the Pribnow box). σ702also forms crystals that are suitable for X-ray analysis. σ703 – 4bound the T4 AsiA protein with high affinity. The epitope for T4 AsiA on σ70was further localized to within σ70[551 – 608], comprising σ conserved region 4.2.