Backbone dynamics of the C-terminal domain of Escherichia coli topoisomerase I in the absence and presence of single-stranded DNA

The backbone dynamics of the C-terminal DNA-binding domain of Escherichia coli topoisomerase I has been characterized in the absence and presence of single-stranded DNA by NMR spectroscopy. N-15 spin-lattice relaxation times (T-1), spin-spin relaxation times (T-2), and heteronuclear NOEs were determined for the uniformly N-15-labeled protein. These data were analyzed by using the model-free formalism to derive the model-free parameters (S-2, tau(e), and R(ex)) for each backbone N-H bond vector and the overall molecular rotational correlation time (tau(m)), The molecular rotational correlation time tau(m) was determined to be 7.49 +/- 0.36 ns for the free and 12.7 +/- 1.07 ns for the complexed protein. Several residues were found to be much more mobile than the average, including 11 residues at the N-terminus, 2 residues at the C-terminus, and residues 25 and 31-35 which are located in a region of the protein that binds to DNA. The binding of ssDNA to the free protein causes a slight increase in the order parameters (S-2) for a small number of residues and a slight decrease in the order parameters (S-2) for the majority of the residues. In particular, upon binding to ssDNA, the mobility of the first alpha-helix and the two beta-sheets was slightly increased, and the mobility of a few specific residues in the loops/turns was restricted. These results differ from the previous studies on the backbone dynamics of molecular complexes in which reduced mobilities were typically observed upon ligand binding.