Figure 3.

Promoter complexes engaged in abortive RNA synthesis undergo a temperature-dependent conformational change. (A) Promoter complexes were formed at 37°C in the presence or absence of 0.5 mM CpG and 50 μM CTP, incubated for 15 min, and footprinted as described in the legend of Fig. 1. (B) KMnO₄ probing of promoter complexes formed at 37°C in the presence of CpG (0.5 mM) and different individual NTPs (50 μM each). (C) KMnO₄ probing of promoter complexes formed at low temperature in the presence of CpG and CTP. Promoter complexes were formed on ice or at 37°C and incubated for 15 min. Reactions shown in lanes 2–4 and 6–8 were supplemented with 0.5 mM CpG and 50 μM CTP, incubated for 15 min, and probed with KMnO₄ (lanes 2, 4, 6, and 8). Complexes shown in lanes 3 and 7 were formed at 37°C, incubated in the presence of transcription substrates, transferred on ice, and incubated for an additional 15 min before KMnO₄ probing. Lanes 1 and 5 show complexes formed in the absence of transcription substrates at 37°C.