Figure 4.
Coupled expression and detection of Ec RNAP mutants in the reconstituted protein synthesis system. The Fluc activity was measured in protein synthesis reactions containing the linear DNA templates of dnaKp1-Fluc and PT7-σ32, separate plasmid templates encoding all four or three wild-type Ec RNAP subunits (PT7-αββ′ω or PT7-αβ′ω) and the PCR-generated linear template for the wild-type β subunit [PT7-β (WT)] or the single-substitution β subunit mutant [PT7-β (Q513P), PT7-β (Q513L) or PT7-β (H526Y)] in the absence or presence of rifampicin (1.2 μM). The relative Fluc activity was presented with the activity from the template of PT7-αββ′ω set as 100. All data were obtained from at least two independent protein synthesis reactions.