title: Structure and DNA-binding properties of the cytolysin regulator CylR2 from Enterococcus faecalis.
abstract: Enterococcus faecalis is one of the major causes for hospital-acquired antibiotic-resistant infections. It produces an exotoxin, called cytolysin, which is lethal for a wide range of Gram-positive bacteria and is toxic to higher organisms. Recently, the regulation of the cytolysin operon was connected to autoinduction by a quorum-sensing mechanism involving the CylR1/CylR2 two-component regulatory system. We report here the crystal structure of CylR2 and its properties in solution as determined by heteronuclear NMR spectroscopy. The structure reveals a rigid dimer containing a helix-turn-helix DNA-binding motif as part of a five-helix bundle that is extended by an antiparallel beta-sheet. We show that CylR2 is a DNA-binding protein that binds specifically to a 22 bp fragment of the cytolysin promoter region. NMR chemical shift perturbation experiments identify surfaces involved in DNA binding and are in agreement with a model for the CylR2/DNA complex that attributes binding specificity to a complex network of CylR2/DNA interactions. Our results propose a mechanism where repression is achieved by CylR2 obstruction of the promoter preventing biosynthesis of the cytolysin operon transcript.
authors.affiliation: Department for NMR-based Structural Biology, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany.
authors.forename: Sigrun
authors.initials: S
authors.lastname: Rumpel
authors.forename: Adelia
authors.initials: A
authors.lastname: Razeto
authors.forename: Chris M
authors.initials: CM
authors.lastname: Pillar
authors.forename: Vinesh
authors.initials: V
authors.lastname: Vijayan
authors.forename: Austin
authors.initials: A
authors.lastname: Taylor
authors.forename: Karin
authors.initials: K
authors.lastname: Giller
authors.forename: Michael S
authors.initials: MS
authors.lastname: Gilmore
authors.forename: Stefan
authors.initials: S
authors.lastname: Becker
authors.forename: Markus
authors.initials: M
authors.lastname: Zweckstetter
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D001430:Bacteriocins; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D019281:Dimerization; D013293:Enterococcus faecalis / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D019077:Helix-Turn-Helix Motifs; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D009876:Operon; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D012097:Repressor Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D001430:Bacteriocins; D004268:DNA-Binding Proteins; D012097:Repressor Proteins; C120351:cytolysin, Enterococcus faecalis