Journal
NUCLEIC ACIDS RESEARCH
Volume 41, Issue 21, Pages 9945-9955Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkt769
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Funding
- Wellcome Trust International Senior Research Fellowship [081760]
- Foundation for Polish Science 'Ideas for Poland'
- Polish Ministry of Science and Higher Education [ESRF/73/2006]
- European Community's Seventh Framework Program [226716]
- Wellcome Trust Senior Research Fellowship
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The key intermediate in genetic recombination is the Holliday junction (HJ), a four-way DNA structure. At the end of recombination, HJs are cleaved by specific nucleases called resolvases. In Gram-negative bacteria, this cleavage is performed by RuvC, a dimeric endonuclease that belongs to the retroviral integrase superfamily. Here, we report the first crystal structure of RuvC in complex with a synthetic HJ solved at 3.75 A resolution. The junction in the complex is in an unfolded 2-fold symmetrical conformation, in which the four arms point toward the vertices of a tetrahedron. The two scissile phosphates are located one nucleotide from the strand exchange point, and RuvC approaches them from the minor groove side. The key protein-DNA contacts observed in the structure were verified using a thiol-based site-specific cross-linking approach. Compared with known complex structures of the phage resolvases endonuclease I and endonuclease VII, the RuvC structure exhibits striking differences in the mode of substrate binding and location of the cleavage site.
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