Localization of The Initiation Site in T7 RNA Polymerase by Affinity Labeling

The oligonucleotide duplexes containing modified CMP residues with the additional beta-D-ribofuranosyl group were used to study the interactions between T7 RNA polymerase (T7 RNAP) and its promoter. Wild-type enzyme did not undergo the affinity labelling with these reagents. However the use of Y639K T7 RNAP mutant resulted in the specific covalent binding thus suggesting the location of the a.a. residue in pos. 639 within the transcription initiation site.