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 1 H– 15 N correlation spectra of a 15 N-labelled RTP.C110S mutant upon the addition of a 21 base pair symmetrical DNA binding site. Assignment of the 1 H– 15 N correlations was achieved using a suite of triple resonance NMR experiments with 15 N, 13 C,70% 2 H enriched protein recorded at 800 MHz and using TROSY pulse sequences. Perturbations to 1 H– 15 N spectra revealed that the N-termini, α3-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.