Substitution of Ribonucleotides in the T7 RNA Polymerase Promoter Element

Substitution of ribonucleotides in the T7 RNA polymerase promoter element.

A systematic analysis was carried out to examine the effects of ribonucleotide substitution at various locations within the promoter element for T7 RNA polymerase. Ribonucleotides could be introduced at most positions without significantly decreasing transcription efficiency. A critical window of residues that were intolerant of RNA substitution was defined for both the nontemplate and template...

Promoter specificity determinants of T7 RNA polymerase.

The high specificity of T7 RNA polymerase (RNAP) for its promoter sequence is mediated, in part, by a specificity loop (residues 742-773) that projects into the DNA binding cleft (1). Previous work demonstrated a role for the amino acid residue at position 748 (N748) in this loop in discrimination of the base pairs (bp) at positions -10 and -11 (2). A comparison of the sequences of other phage ...

Identification of a minimal binding element within the T7 RNA polymerase promoter.

The T7 RNA polymerase promoter has been proposed to contain two domains: the binding region upstream of position -5 is recognized through apparently traditional duplex contacts, while the catalytic domain downstream of position -5 is bound in a melted configuration. This model is tested by following polymerase binding to a series of synthetic oligonucleotides representing truncations of the con...

Evidence for DNA bending at the T7 RNA polymerase promoter.

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 studies 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 its promoter DNA. A com...

Structure and function in promoter escape by T7 RNA polymerase.