Abstract

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 a 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 15 N-labeling of the protein and processes unassigned H N- 15 N HSQC spectra, H N- 15 N RDCs, and sparse H N-H N 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 (H N, 15 N) backbone resonances as well as the dNNs of the 3D 15 N-NOESY spectrum, in O(n 3) time. The algorithm is demonstrated on NMR data from a 76-residue protein, human ubiquitin, matched to four structures, including

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