Journal article
Fork-remodeling helicase Rad5 preferentially reverses replication forks with gaps in the leading strand
Journal of molecular biology, Vol.435(4), 167946
01/06/2023
DOI: 10.1016/j.jmb.2023.167946
PMCID: PMC9915103
PMID: 36623584
Abstract
DNA damage bypass pathways promote the replication of damaged DNA when replication forks stall at sites of DNA damage. Template switching is a DNA damage bypass pathway in which fork-reversal helicases convert stalled replication forks into four-way DNA junctions called chicken foot intermediates, which are subsequently extended by replicative DNA polymerases. In yeast, fork-reversal is carried out by the Rad5 helicase using an unknown mechanism. To better understand the mechanism of Rad5 and its specificity for different fork DNA substrates, we used a FRET-based assay to observe fork reversal in real time. We examined the ability of Rad5 to bind and catalyze the reversal of various fork DNA substrates in the presence of short gaps in the leading or lagging strand as well as in the presence or absence of RPA and RNA primers in the lagging strand. We found that Rad5 preferentially reverses fork DNA substrates with short gaps (10 to 30 nt.) in the leading strand. Thus, Rad5 preferentially reverses fork DNA substrates that form chicken foot intermediates with 5' overhangs that can be extended by replicative DNA polymerases during the subsequent steps of template switching.
Details
- Title: Subtitle
- Fork-remodeling helicase Rad5 preferentially reverses replication forks with gaps in the leading strand
- Creators
- Justin A Ling - University of IowaMelissa S Gildenberg - University of IowaMasayoshi Honda - University of IowaChristine M Kondratick - University of IowaMaria Spies - University of IowaM Todd Washington - Department of Biochemistry, University of Iowa College of Medicine, Iowa City, IA 52242-1109. Electronic address: todd-washington@uiowa.edu
- Resource Type
- Journal article
- Publication Details
- Journal of molecular biology, Vol.435(4), 167946
- DOI
- 10.1016/j.jmb.2023.167946
- PMID
- 36623584
- PMCID
- PMC9915103
- NLM abbreviation
- J Mol Biol
- ISSN
- 0022-2836
- eISSN
- 1089-8638
- Grant note
- DOI: 10.13039/100000002, name: National Institutes of Health; DOI: 10.13039/100000054, name: National Cancer Institute, award: CA232425; DOI: 10.13039/100000057, name: National Institute of General Medical Sciences, award: GM081433
- Language
- English
- Date published
- 01/06/2023
- Academic Unit
- Radiation Oncology; Biochemistry and Molecular Biology
- Record Identifier
- 9984357393702771
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