Interactions in the active site of Deinococcus radiodurans RNA polymerase during RNA proofreading.
Co-transcriptional RNA proofreading by RNA polymerase (RNAP) is essential for accurate mRNA synthesis and reactivation of stalled transcription complexes, which can otherwise compromise genome integrity. RNAP from the stress-resistant bacterium Deinococcus radiodurans exhibits high levels of RNA cleavage in comparison with RNAP from Escherichia coli, which allows it to remove misincorporated nucleotides with high efficiency. Here, we show that the rate of RNA cleavage by D. radiodurans RNAP depends on the structure of the (mis)matched RNA 3'-nucleotide and its contacts with the active site. These interactions likely position the reactive phosphodiester bond in the cleavage-competent conformation, thus facilitating its hydrolysis catalyzed by metal ions in the active center. The universal RNA cleavage factor GreA largely alleviates defects in RNA cleavage caused by modifications in the RNA 3'-nucleotide or in its binding pocket in RNAP, suggesting that GreA functionally substitutes for these contacts. The results demonstrate that various RNAPs rely on a conserved mechanism for RNA proofreading, which can be modulated by changes in accessory parts of the active center.
['Esyunina|Daria|D|', 'Kulbachinskiy|Andrey|A|']
D001426:Bacterial Proteins D020134:Catalytic Domain D012321:DNA-Directed RNA Polymerases D034301:Deinococcus D008345:Manganese D008958:Models, Molecular D009690:Nucleic Acid Conformation D059367:RNA Cleavage D012329:RNA, Bacterial
Deinococcus radiodurans RNA polymerase RNA proofreading GreA Stress resistance