Mapping protein dynamics conformational changes in replisomal macromolecular assemblies
Understanding how proteins interact and behave in large dynamic multi-protein complexes is an important area of significance in many human diseases that are associated with faults in their interactions. Increasing understanding of the well characterized bacterial DNA replication machinery (the replisome) will continue to enrich our understanding of other dynamic complexes, including those that carry out human DNA replication. Although replication proteins are not themselves highly conserved between eubacteria and higher organisms, the replication mechanisms generally are. We focussed on pairwise interactions that involve one of the key organizational centres of the bacterial replisome, the DnaI helicase loader and the DnaB helicase. As a first step, we measured the dynamics of the Bacillus sp. helicase loader DnaI and the DnaB helicase separately, using elastic, quasielastic and inelastic neutron scattering at four instrumental resolutions. This provides a basis for future experiments on the DnaI-DnaB complex. The measurement established that internal molecular flexibility and diffusive motions are significantly higher in DnaB helicase compared to those evaluated in DnaI. The structural rigidity was also studied. The analysis revealed that DnaI structural rigidity is larger than that of DnaB. Furthermore, the method allowed the extraction of vibrational density of states, showing that DnaI has more vibrational modes than DnaB.