title: Structural analysis of the chromosome segregation protein Spo0J from Thermus thermophilus.
abstract: Prokaryotic chromosomes and plasmids encode partitioning systems that are required for DNA segregation at cell division. The plasmid partitioning loci encode two proteins, ParA and ParB, and a cis-acting centromere-like site denoted parS. The chromosomally encoded homologues of ParA and ParB, Soj and Spo0J, play an active role in chromosome segregation during bacterial cell division and sporulation. Spo0J is a DNA-binding protein that binds to parS sites in vivo. We have solved the X-ray crystal structure of a C-terminally truncated Spo0J (amino acids 1-222) from Thermus thermophilus to 2.3 A resolution by multiwavelength anomalous dispersion. It is a DNA-binding protein with structural similarity to the helix-turn-helix (HTH) motif of the lambda repressor DNA-binding domain. The crystal structure is an antiparallel dimer with the recognition alpha-helices of the HTH motifs of each monomer separated by a distance of 34 A corresponding to the length of the helical repeat of B-DNA. Sedimentation velocity and equilibrium ultracentrifugation studies show that full-length Spo0J exists in a monomer-dimer equilibrium in solution and that Spo0J1-222 is exclusively monomeric. Sedimentation of the C-terminal domain of Spo0J shows it to be exclusively dimeric, confirming that the C-terminus is the primary dimerization domain. We hypothesize that the C-terminus mediates dimerization of Spo0J, thereby effectively increasing the local concentration of the N-termini, which most probably dimerize, as shown by our structure, upon binding to a cognate parS site.
authors.affiliation: MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK. tleonard@mrc-lmb.cam.ac.uk
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Leonard
authors.forename: P Jonathan G
authors.initials: PJ
authors.lastname: Butler
authors.forename: Jan
authors.initials: J
authors.lastname: Löwe
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D020090:Chromosome Segregation; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D019281:Dimerization; D019077:Helix-Turn-Helix Motifs; D008958:Models, Molecular; D008970:Molecular Weight; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D016963:Thermus thermophilus / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D004247:DNA