E. Severinova et al., DOMAIN ORGANIZATION OF THE ESCHERICHIA-COLI RNA-POLYMERASE SIGMA(70) SUBUNIT, Journal of Molecular Biology, 263(5), 1996, pp. 637-647
We used limited trypsin digestion to determine the domain organization
of the Escherichia coli RNA polymerase sigma(70) subunit. Trypsin-res
istant fragments containing sigma(70) conserved region 2 (sigma(2)(70)
), and carboxy-terminal fragments containing conserved regions 3 and 4
(sigma(3-4)(70)) were identified by a combination of amino acid seque
ncing and mass spectrometry. The domains were studied for partial bioc
hemical functions of sigma(70).sigma(2)(70) bound core RNA polymerase
competitively with intact sigma(70). In contrast to sigma(2)(70) alone
, the RNA polymerase holoenzyme formed with sigma(2)(70) 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)(70) also forms crystals that are suitable for X-ray analysis
. sigma(3-4)(70) bound the T4 AsiA protein with high affinity. The epi
tope for T4 AsiA on sigma(70) was further localized to within sigma(70
)[551-608], comprising sigma conserved region 4.2. (C) 1996 Academic P
ress Limited