title: Crystal structure of RuvC resolvase in complex with Holliday junction substrate.
abstract: 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 Å 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.
authors: Laboratory of Protein Structure, International Institute of Molecular and Cell Biology, 4 Trojdena Street, 02-109, Warsaw, Poland.
authors: Karolina M
authors: KM
authors: Górecka
authors: Weronika
authors: W
authors: Komorowska
authors: Marcin
authors: M
authors: Nowotny
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D045566:DNA, Cruciform / Q000737:chemistry* / Q000378:metabolism; D045565:Holliday Junction Resolvases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data
chemical_list: D001426:Bacterial Proteins; D045566:DNA, Cruciform; D045565:Holliday Junction Resolvases
references: Biochemistry. 1999 Aug 31;38(35):11349-58
references: 10471285
references: Genes Cells. 1998 Sep;3(9):575-86
references: 9813108
references: J Mol Biol. 2001 Nov 2;313(4):751-64
references: 11697901
references: Structure. 2001 Mar 7;9(3):197-204
references: 11286886
references: Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6063-7
references: 1829835
references: Genet Res. 2007 Dec;89(5-6):285-307
references: 18976517
references: Nature. 2008 Nov 20;456(7220):357-61
references: 19020614
references: Mol Microbiol. 2001 Feb;39(4):823-34
references: 11251805
references: J Mol Biol. 1995 Sep 15;252(2):213-26
references: 7674302
references: Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32
references: 15572765
references: Acta Crystallogr D Biol Crystallogr. 2010 Feb;66(Pt 2):125-32
references: 20124692
references: Nature. 2007 Oct 4;449(7162):621-4
references: 17873858
references: Biochemistry. 2000 Oct 3;39(39):11982-8
references: 11009612
references: Acta Crystallogr D Biol Crystallogr. 1999 Apr;55(Pt 4):849-61
references: 10089316
references: Nucleic Acids Res. 2006;34(19):5577-84
references: 17028102
references: J Synchrotron Radiat. 2012 May;19(Pt 3):442-9
references: 22514183
references: Nature. 2007 Oct 4;449(7162):616-20
references: 17873859
references: Science. 1998 Nov 27;282(5394):1669-75
references: 9831551
references: J Biol Chem. 1996 Oct 18;271(42):26105-9
references: 8824253
references: Mol Microbiol. 2013 Sep;89(6):1240-58
references: 23888987
references: Cell. 1988 Oct 7;55(1):79-89
references: 3167979
references: Annu Rev Biochem. 1992;61:603-40
references: 1497320
references: Cell. 1994 Sep 23;78(6):1063-72
references: 7923356
references: EMBO J. 1991 Dec;10(13):4381-9
references: 1661673
references: Acta Crystallogr D Biol Crystallogr. 1998 Sep 1;54(Pt 5):905-21
references: 9757107
references: Genes Dev. 2008 Apr 15;22(8):1093-106
references: 18413719
references: J Bacteriol. 2010 Oct;192(20):5499-505
references: 20729360
references: J Mol Biol. 1998 Apr 24;278(1):117-33
references: 9571038
references: Genes Cells. 2000 Oct;5(10):803-13
references: 11029656
references: Mol Cell. 2002 Sep;10(3):671-81
references: 12408833
references: Nucleic Acids Res. 1998 Apr 1;26(7):1560-6
references: 9512524
references: Science. 2006 Feb 24;311(5764):1153-7
references: 16497933
references: Cell. 1994 Dec 2;79(5):853-64
references: 8001122
references: Acta Crystallogr D Biol Crystallogr. 2010 Feb;66(Pt 2):213-21
references: 20124702
references: Q Rev Biophys. 2000 May;33(2):109-59
references: 11131562
references: J Biol Chem. 2001 Mar 30;276(13):10432-6
references: 11152689
references: Nature. 1991 Dec 19-26;354(6354):506-10
references: 1758493
references: Nucleic Acids Res. 2013 Jan 7;41(1):648-56
references: 23118486
references: BMC Genomics. 2006 Apr 28;7:98
references: 16646971
references: Proc Natl Acad Sci U S A. 2001 May 8;98(10):5509-14
references: 11331763
references: J Appl Crystallogr. 2007 Aug 1;40(Pt 4):658-674
references: 19461840
references: EMBO J. 1990 May;9(5):1665-72
references: 2184035
references: Cell. 1993 Sep 24;74(6):1021-31
references: 8402879
references: EMBO Rep. 2009 Feb;10(2):144-51
references: 19165139
references: Proc Natl Acad Sci U S A. 1995 Aug 1;92(16):7470-4
references: 7638215