A1 Refereed original research article in a scientific journal
Role of the trigger loop in translesion RNA synthesis by bacterial RNA polymerase
Authors: Aleksei Agapov, Artem Ignatov, Matti Turtola, Georgiy Belogurov, Daria Esyunina, Andrey Kulbachinskiy
Publisher: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Publication year: 2020
Journal: Journal of Biological Chemistry
Journal name in source: JOURNAL OF BIOLOGICAL CHEMISTRY
Journal acronym: J BIOL CHEM
Volume: 295
Issue: 28
First page : 9583
Last page: 9595
Number of pages: 13
ISSN: 0021-9258
DOI: https://doi.org/10.1074/jbc.RA119.011844
Self-archived copy’s web address: https://research.utu.fi/converis/portal/detail/Publication/49774259
DNA lesions can severely compromise transcription and block RNA synthesis by RNA polymerase (RNAP), leading to subsequent recruitment of DNA repair factors to the stalled transcription complex. Recent structural studies have uncovered molecular interactions of several DNA lesions within the transcription elongation complex. However, little is known about the role of key elements of the RNAP active site in translesion transcription. Here, using recombinantly expressed proteins,in vitrotranscription, kinetic analyses, andin vivocell viability assays, we report that point amino acid substitutions in the trigger loop, a flexible element of the active site involved in nucleotide addition, can stimulate translesion RNA synthesis byEscherichia coliRNAP without altering the fidelity of nucleotide incorporation. We show that these substitutions also decrease transcriptional pausing and strongly affect the nucleotide addition cycle of RNAP by increasing the rate of nucleotide addition but also decreasing the rate of translocation. The secondary channel factors DksA and GreA modulated translesion transcription by RNAP, depending on changes in the trigger loop structure. We observed that although the mutant RNAPs stimulate translesion synthesis, their expression is toxicin vivo, especially under stress conditions. We conclude that the efficiency of translesion transcription can be significantly modulated by mutations affecting the conformational dynamics of the active site of RNAP, with potential effects on cellular stress responses and survival.
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