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- Title
Visualization of human Bloom's syndrome helicase molecules bound to homologous recombination intermediates.
- Authors
Gyimesi, Máté; Pires, Ricardo H.; Billington, Neil; Sarlós, Kata; Kocsis, Zsuzsa S.; Módos, Károly; Kellermayer, Miklós S. Z.; Kovács, Mihály
- Abstract
Homologous recombination (HR) is a key process in the repair of double-stranded DNA breaks (DSBs) that can initiate cancer or cell death. Human Bloom's syndrome RecQ-family DNA helicase (BLM) exerts complex activities to promote DSB repair while avoiding illegitimate HR. The oligomeric assembly state of BLM has been a key unresolved aspect of its activities. In this study we assessed the structure and oligomeric state of BLM, in the absence and presence of key HR-intermediate DNA structures, by using single-molecule visualization (electron microscopic and atomic force microscopic single-particle analysis) and solution biophysical (dynamic light scattering, kinetic and equilibrium binding) techniques. Besides full-length BLM, we used a previously characterized truncated construct (BLM642-1290) as a monomeric control. Contrary to previous models proposing a ring-forming oligomer, we found the majority of BLM molecules to be monomeric in all examined conditions. However, BLM showed a tendency to form dimers when bound to branched HR intermediates. Our results suggest that HR activities requiring single-stranded DNA translocadon are performed by monomeric BLM, while complex DNA structures encountered and dissolved by BLM in later stages of HR induce partial oligomerization of the helicase.
- Subjects
HELICASES; ENZYMES; MOLECULES; MOLECULAR physics; DNA
- Publication
FASEB Journal, 2013, Vol 27, Issue 12, p4954
- ISSN
0892-6638
- Publication type
Article
- DOI
10.1096/fj.13-234088