Figure 2. T7 gp2 inhibits RNAP interaction with the T5 N25 promoter.

(a) Time dependence of the change in fluorescence upon mixing 1 nM [211Cys-TMR] σ⁷⁰ RNAP holoenzyme with 2 nM −65/+35 T5N25 (black) or a non-promoter DNA fragment (red) in the presence or in the absence of gp2. The upper curve (labeled as “(RNAP+T5N25)+gp2” was obtained in experiment where gp2 was added to the RNAP beacon preincubated with 2 nM T5N25 promoter fragment for 30 min. In other experiments, gp2 was added to preformed RNAP-T5N25 or RNAP-non-promoter DNA complexes.

(b) Inhibition of abortive transcript synthesis from T5N25 promoter fragment by gp2. Reactions contained Eσ⁷⁰ alone (lane 1) or Eσ⁷⁰ with gp2 added before (lane 2) or after (lane 3) open complex formation. An autoradiograph of denaturing polyacrylamide gel is presented.