FIG 2.

The relatively conserved R2 region is essential for the function of M. tuberculosis ω. (A) Sequence alignment of the ω subunits from M. tuberculosis, E. coli, and T. thermophilus. The amino acid numbering shown on top of the this figure is for M. tuberculosis ω. (B) SDS-PAGE analysis of purified M. tuberculosis coreΔω coexpressed with Flag-tagged ω from M. tuberculosis, E. coli, or T. thermophilus. Western blots of ω^Flag with a Flag tag antibody are shown in the bottom panel. (C) Purified M. tuberculosis core with a mutated ω subunit. M. tuberculosis ω mutants, in which the R1, R2, or R3 regions of M. tuberculosis ω was replaced with the corresponding regions of E. coli ω (ω^E-R1, ω^E-R2, and ω^E-R3), respectively, were coexpressed with M. tuberculosis coreΔω in E. coli. Western blots of mutated ω^Flag subunit with a Flag tag antibody are shown in the bottom panel. (D) In vitro abortive transcription of purified M. tuberculosis coreΔω derivatives at Rv1494p. RbpA and σ^A were added in each reaction in this promoter-dependent transcription assay. A representative result from three independent tests is shown. Mtb, M. tuberculosis; Eco, E. coli; Tth, T. thermophilus.