Abstract
We used limited trypsin digestion to determine the domain organization of the Escherichia coli RNA polymerase sigma subunit. Trypsin-resistant fragments containing sigma conserved region 2 (sigma 2 ), and carboxy-terminal fragments containing conserved regions 3 and 4 (sigma 3 - 4 ) were identified by a combination of amino acid sequencing and mass spectrometry. The domains were studied for partial biochemical functions of sigma ?sigma 2 bound core RNA polymerase competitively with intact sigma . In contrast to sigma 2 alone, the RNA polymerase holoenzyme formed with sigma 2 specifically bound a single-stranded DNA oligomer with a sequence corresponding to the non-template strand of the -10 promoter element (the Pribnow box). sigma 2 also forms crystals that are suitable for X-ray analysis. sigma 3 - 4 bound the T4 AsiA protein with high affinity. The epitope for T4 AsiA on sigma was further localized to within sigma [551 - 608], comprising sigma conserved region 4.2.