Fig. 2.

Transcriptional activities of polymerases reconstituted with mutant σ proteins using P_uvsX in vitro in the presence of AsiA and/or MotA. The indicated σ (0.2 pmol) and AsiAHis₆ (3 pmol ) were incubated in 1.8 μl of protein buffer I for 10 min at 37° C. A solution containing 0.05 pmol of core in 1.1 μl of protein buffer II was then added, and the resulting solution was incubated for an additional 10 min at 37° C. Transcription was initiated by adding this solution to 2.1 μl of DNA buffer containing 0.01 pmol DNA. After 37° C for 20 sec, rifampicin (150 ng) was added to limit transcription to a single round. For each indicated σ, [the amount of P_uvsX RNA observed]/[the amount of P_usvX RNA observed with σ⁷⁰ polymerase in the absence of MotA and AsiA] is shown using polymerase alone, polymerase with MotA, polymerase with AsiA, and polymerase with AsiA and MotA. Values were determined from two or more transcriptions. Error is indicated by darker shaded area. In some cases where the level of transcription was low, 10-fold values (10 × values) are also shown. In these cases, zero is the bottom of the enclosed box.