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Altered Minor‐Groove Hydrogen Bonds in DNA Block Transcription Elongation by T7 RNA Polymerase

DNA transcription depends upon the highly efficient and selective function of RNA polymerases (RNAPs). Modifications in the template DNA can impact the progression of RNA synthesis, and a number of DNA adducts, as well as abasic sites, arrest or stall transcription. Nonetheless, data are needed to understand why certain modifications to the structure of DNA bases stall RNA polymerases while others are efficiently bypassed. In this study, we evaluate the impact that alterations in dNTP/rNTP base‐pair geometry have on transcription. T7 RNA polymerase was used to study transcription over modified purines and pyrimidines with altered H‐bonding capacities. The results suggest that introducing wobble base‐pairs into the DNA:RNA heteroduplex interferes with transcriptional elongation and stalls RNA polymerase. However, transcriptional stalling is not observed if mismatched base‐pairs do not H‐bond. Together, these studies show that RNAP is able to discriminate mismatches resulting in wobble base‐pairs, and suggest that, in cases of modifications with minor steric impact, DNA:RNA heteroduplex geometry could serve as a controlling factor for initiating transcription‐coupled DNA repair.