Date of Original Version
Abstract or Description
High-throughput NMR structural biology can play an important role in structural genomics. We report an automated procedure for high-throughput NMR resonance assignment for a protein of known structure, or of an homologous structure. These assignments are a prerequisite for probing protein-protein interactions, protein-ligand binding, and dynamics by NMR. Assignments are also the starting point for structure determination and refinement. A new algorithm, called Nuclear Vector Replacement (NVR) is introduced to compute assignments that optimally correlate experimentally-measured NH residual dipolar couplings (RDCs) to a given a priori whole-protein 3D structural model. The algorithm requires only uniform 15N-labelling of the protein, and processes unassigned HN-15N HSQC spectra, HN-15N RDCs, and sparse HN-HN NOE's dNNs), all of which can be acquired in a fraction of the time needed to record the traditional suite of experiments used to perform resonance assignments. NVR runs in minutes and efficiently assigns the (HN,15N) backbone resonances as well as the dNNs of the 3D \nfif-NOESY spectrum, in O(n3) time. The algorithm is demonstrated on NMR data from a 76-residue protein, human ubiquitin, matched to four structures, including one mutant (homolog), determined either by X-ray crystallography or by different NMR experiments (without RDCs). NVR achieves an average assignment accuracy of over 90%. We further demonstrate the feasibility of our algorithm for different and larger proteins, using NMR data for hen lysozyme (129 residues, 98% accuracy) and streptococcal protein G (56 residues, 95% accuracy), matched to a variety of 3D structural models. Finally, we extend NVR to a second application, 3D structural homology detection, and demonstrate that NVR is able to identify structural homologies between proteins with remote amino acid sequences using a database of structural models.
Proceedings of The Seventh Annual International Conference on Computational Molecular Biology (RECOMB), 176-187.