Nuclear magnetic resonance analysis of protein-DNA interactions

Abstract

Recent methodological and instrumental advances in solution-state nuclear magnetic resonance have opened up the way to investigating challenging problems in structural biology such as large macromolecular complexes. This review focuses on the experimental strategies currently employed to solve structures of protein-DNA complexes and to analyse their dynamics. It highlights how these approaches can help in understanding detailed molecular mechanisms of target recognition.

Figure 1.

Interaction surface mapping by combining chemical shift perturbation (CSP) and imino cross saturation on the THAP domain of hTHAP1. (a) Histogram of the normalized CSP observed upon DNA binding as a function of the residue number. (b) Imino cross-saturation rates (R_D) as a function of residue number. (c) Examples of experimental points and fitted curves of the imino cross-saturation data. Experimental points and fitted curves are coloured in blue for the α-helical residues (away from DNA) and in black for β-sheet residues (close to DNA). (d) Mapping of the interaction surface on the solution structure of the THAP domain of hTHAP1.

Figure 2.

The residual dipolar coupling formalism, illustration and equations.

Figure 3.

Long-range distance restraints from PRE, adapted from Iwahara et al. [58]. Impact of intermolecular PRE data on coordinate accuracy of the SRY–DNA complex when only a single intermolecular NOE restraint located at the centre of the protein–DNA interface is employed. Best-fit superposition of 30 simulated annealing structures (SRY, red; DNA, blue) calculated (a) without and (b) with 438 intermolecular ¹H-PRE restraints.

Figure 4.

Computational strategies for calculation of protein–DNA complex structures using a data-driven docking protocol. (a) Modelling of the complex formed by the THAP domain of hTHAP1 and its DNA target, including solely interface mapping restraints as ambiguous interface restraints. On the graph, the intermolecular energy is plotted as a function of RMSD from the lowest energy structure in order to illustrate the structure calculation convergence. On the right-hand side, a superposition of the 15 best models is shown. (b) Modelling of the complex formed by the THAP domain of hTHAP1 and its DNA target, including interface mapping restraints as ambiguous interface restraints, 39 intermolecular NOEs and 49 ¹D_HN-N RDCs for refinement. In the graph, the intermolecular energy is plotted as a function of RMSD from the lowest energy structure in order to illustrate the calculation convergence. On the right-hand side, a superposition of the 15 best structures is shown (PDB ID 2ko0). (c) Modelling of the complex formed by the THAP domain of hTHAP1 and its DNA target, including interface mapping restraints as ambiguous interface restraints, 39 intermolecular NOEs and 49 ¹D_HN-N RDCs and 49 ¹D_C′-Cα RDCs for refinement. In the graph, intermolecular energy is plotted as a function of RMSD from the lowest energy structure in order to illustrate the calculation convergence. On the right-hand side, a superposition of the 15 best structures is shown and, in both graphs, a comparison between experimentally measured RDCs and backcalculated RDCs is shown for ¹D_HN-N RDCs and ¹D_C′-Cα RDCs. For each ensemble, pair-wise RMSD on backbone heavy atoms is shown.