Phage T7 RNA polymerase is the only DNA-dependent RNA polymerase for which
we have a high-resolution structure of the promoter-bound complex. Recent s
tudies with the more complex RNA polymerases have suggested a role for DNA
wrapping in the initiation of transcription. Here, circular permutation gel
retardation assays provide evidence that the polymerase does indeed bend i
ts promoter DNA. A complementary set of experiments employing differential
phasing from an array of phased A-tracts provides further evidence for both
intrinsic and polymerase-induced bends in the T7 RNA polymerase promoter D
NA. The bend in the complex is predicted to be about 40-60 degrees and to b
e centered around positions -2 to +1, at the start site for transcription,
while the intrinsic bend is much smaller (about 10 degrees). These results,
viewed in the light of a recent crystal structure for the complex, suggest
a mechanism by which binding leads directly to bending. Bending at the sta
rt site would then facilitate the melting necessary to initiate transcripti
on. (C) 2000 Academic Press.