T7 RNA-POLYMERASE MUTANTS WITH ALTERED PROMOTER SPECIFICITIES

The amino acid at position 748 in T7 RNA polymerase (RNAP) functions t o discriminate base pairs at positions -10 and -11 in the promoter. We have constructed a series of T7 RNAP mutants having all possible amin o acid substitutions at this position. Surprisingly, most (13/19) subs titutions result in active RNAPs, and many of these exhibit altered pr omoter specificities. Identification of mutant RNAPs with altered spec ificities expands the repertoire of highly specific phage RNAPs that a re available for use in phage RNAP-based transcription systems and hig hlights the complexity of sequence-specific DNA recognition.