1. The unstructured linker arms of MutL enable GATC site incision beyond roadblocks during initiation of DNA mismatch repair
- Author
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Roland Kanaar, Enes Sahin, Katerina Nitsenko, Peter Friedhoff, Audrey Quessada-Vial, Terence R. Strick, Herrie H.K. Winterwerp, Camille Duboc, Charlie Laffeber, Yannicka S N Mardenborough, Titia K. Sixma, Joyce H.G. Lebbink, Molecular Genetics, and Radiation Oncology
- Subjects
DNA, Bacterial ,Base Pair Mismatch ,Genome Integrity, Repair and Replication ,medicine.disease_cause ,DNA Mismatch Repair ,Genomic Instability ,03 medical and health sciences ,Endonuclease ,chemistry.chemical_compound ,0302 clinical medicine ,CRISPR-Associated Protein 9 ,RNA polymerase ,Escherichia coli ,Genetics ,medicine ,Deoxyribonucleases, Type II Site-Specific ,030304 developmental biology ,0303 health sciences ,Endodeoxyribonucleases ,biology ,Cas9 ,Escherichia coli Proteins ,030302 biochemistry & molecular biology ,DNA replication ,DNA-Directed RNA Polymerases ,MutS DNA Mismatch-Binding Protein ,Cell biology ,DNA-Binding Proteins ,DNA Repair Enzymes ,MutL Proteins ,chemistry ,biology.protein ,DNA mismatch repair ,Linker ,030217 neurology & neurosurgery ,DNA - Abstract
DNA mismatch repair (MMR) maintains genome stability through repair of DNA replication errors. InEscherichia coli, initiation of MMR involves recognition of the mismatch by MutS, recruitment of MutL, activation of endonuclease MutH and DNA strand incision at a hemimethylated GATC site. Here we studied the mechanism of communication that couples mismatch recognition to daughter strand incision. We investigated the effect of catalytically-deficient Cas9 as well as stalled RNA polymerase as roadblocks placed on DNA in between the mismatch and GATC site in ensemble and single molecule nanomanipulation incision assays. The MMR proteins were observed to incise GATC sites beyond a roadblock, albeit with reduced efficiency. This residual incision is completely abolished upon shortening the disordered linker regions of MutL. These results indicate that roadblock bypass can be fully attributed to the long, disordered linker regions in MutL and establish that communication during MMR initiation occurs along the DNA backbone.
- Published
- 2018
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