title: Interaction of the replication terminator protein of Bacillus subtilis with DNA probed by NMR spectroscopy.
abstract: Termination of DNA replication in Bacillus subtilis involves the polar arrest of replication forks by a specific complex formed between the dimeric 29 kDa replication terminator protein (RTP) and DNA terminator sites. We have used NMR spectroscopy to probe the changes in 1H-15N correlation spectra of a 15N-labelled RTP.C110S mutant upon the addition of a 21 base pair symmetrical DNA binding site. Assignment of the 1H-15N correlations was achieved using a suite of triple resonance NMR experiments with 15N,13C,70% 2H enriched protein recorded at 800 MHz and using TROSY pulse sequences. Perturbations to 1H-15N spectra revealed that the N-termini, alpha3-helices and several loops are affected by the binding interaction. An analysis of this data in light of the crystallographically determined apo- and DNA-bound forms of RTP.C110S revealed that the NMR spectral perturbations correlate more closely to protein structural changes upon complex formation rather than to interactions at the protein-DNA interface.
authors.affiliation: School of Molecular and Microbial Biosciences, University of Sydney, Sydney, NSW 2006, Australia.
authors.forename: Adam F
authors.initials: AF
authors.lastname: Hastings
authors.forename: Gottfried
authors.initials: G
authors.lastname: Otting
authors.forename: Rutger H A
authors.initials: RH
authors.lastname: Folmer
authors.forename: Iain G
authors.initials: IG
authors.lastname: Duggin
authors.forename: R Gerry
authors.initials: RG
authors.lastname: Wake
authors.forename: Matthew C J
authors.initials: MC
authors.lastname: Wilce
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Wilce
mesh_terms: D001412:Bacillus subtilis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000737:chemistry*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008958:Models, Molecular; D009841:Oligonucleotides / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D009841:Oligonucleotides; C063112:rtP protein, Bacillus subtilis; D004247:DNA