Search and processing of Holliday junctions within long DNA by junction-resolving enzymes
More about Open Access at the CrickAuthors list
Artur P Kaczmarczyk Anne-Cécile Déclais Matt Newton Simon Boulton David MJ Lilley David S RuedaAbstract
Resolution of Holliday junctions is a critical intermediate step of homologous recombination in which junctions are processed by junction-resolving endonucleases. Although binding and cleavage are well understood, the question remains how the enzymes locate their substrate within long duplex DNA. Here we track fluorescent dimers of endonuclease I on DNA, presenting the complete single-molecule reaction trajectory for a junction-resolving enzyme finding and cleaving a Holliday junction. We show that the enzyme binds remotely to dsDNA and then undergoes 1D diffusion. Upon encountering a four-way junction, a catalytically-impaired mutant remains bound at that point. An active enzyme, however, cleaves the junction after a few seconds. Quantitative analysis provides a comprehensive description of the facilitated diffusion mechanism. We show that the eukaryotic junction-resolving enzyme GEN1 also undergoes facilitated diffusion on dsDNA until it becomes located at a junction, so that the general resolution trajectory is probably applicable to many junction resolving enzymes.
Journal details
Journal Nature Communications
Volume 13
Issue number 1
Pages 5921
Available online
Publication date
Full text links
Publisher website (DOI) 10.1038/s41467-022-33503-6
Europe PubMed Central 36207294
Pubmed 36207294
Keywords
Related topics
Type of publication