Structural and dynamic studies of DNA recognition by NF-κB p50 RHR homodimer using methyl NMR spectroscopy

Protein dynamics involving higher-energy sparsely populated conformational substates are frequently critical for protein function. This study describes the dynamics of the homodimer (p50)(2) of the p50 Rel homology region (RHR) of the transcription factor NF-kappa B, using C-13 relaxation dispersion experiments with specifically (C-13, H-1)-labeled methyl groups of Ile (delta), Leu and Val. Free (p50)(2) is highly dynamic in solution, showing mu s-ms relaxation dispersion consistent with exchange between the ground state and higher energy substates. These fluctuations propagate from the DNA-binding loops through the core of the domain. The motions are damped in the presence of kappa B DNA, but the NMR spectra of the DNA complexes reveal multiple local conformations of the p50 RHR homodimer bound to certain kappa B DNA sequences. Varying the length and sequence of kappa B DNA revealed two factors that promote a single bound conformation for the complex: the length of the kappa B site in the duplex and a symmetrical sequence of guanine nucleotides at both ends of the recognition motif. The dynamic nature of the DNA-binding loops, together with the multiple bound conformations of p50 RHR with certain kappa B sites, is consistent with variations in the transcriptional activity of the p50 homodimer with different kappa B sequences.

Automated summary

This study investigates the dynamics of the p50 Rel homology region (RHR) homodimer of the transcription factor NF-kappa B. The results show that the homodimer is highly dynamic in solution, with fluctuations propagating from the DNA-binding loops through the core of the domain. The presence of kappa B DNA dampens the motions, but multiple local conformations of the p50 RHR homodimer are observed when bound to certain kappa B DNA sequences. These findings provide insights into the transcriptional activity variations of the p50 homodimer with different kappa B sequences.