Structure determination of a new protein from backbone‐centered NMR data and NMR‐assisted structure prediction

Abstract

Targeting of proteins for structure determination in structural genomic programs often includes the use of threading and fold recognition methods to exclude proteins belonging to well-populated fold families, but such methods can still fail to recognize preexisting folds. The authors illustrate here a method in which limited amounts of structural data are used to improve an initial homology search and the data are subsequently used to produce a structure by data-constrained refinement of an identified structural template. The data used are primarily NMR-based residual dipolar couplings, but they also include additional chemical shift and backbone-nuclear Overhauser effect data. Using this methodology, a backbone structure was efficiently produced for a 10 kDa protein (PF1455) from Pyrococcus furiosus. Its relationship to existing structures and its probable function are discussed. Proteins 2006;65:480-489.

RDC‐assisted modeling of symmetric protein homo‐oligomers

“…The use of RDCs as a source of orientational information has become routine in investigations of protein structure and dynamics (Bax and Grishaev 2005;Prestegard et al 2005). In fact, several groups are working on protocols for solving protein structures using primarily RDC and chemical shift information rather than relying on NOEderived distance restraints (Delaglio et al 2000;Valafar and Prestegard 2003;Prestegard et al 2005;Mayer et al 2006). There have also been two particularly significant applications to the determination of a dimer structure that uses RDCs, which differ from the work presented here in that alignment axes are allowed to float during structure determination and a small number of experimental distance restraints are used to help determine structures (Bewley and Clore 2000;Rumpel et al 2008).…”

Modeling helical proteins using residual dipolar couplings, sparse long-range distance constraints and a simple residue-based force field

“…Here, we have coupled this approach with a minimal force field and Monte Carlo/simulated annealing sampling method to determine the tertiary fold of proteins. As with the approach proposed by Prestegard and coworkers [72], this method does not rely on sequence content and utilized sparse and easily obtainable NMR/EPR data. Unlike the method proposed by Blackledge and coworkers [73], our approach is compatible with the use of dipolar waves derived from only one aligning medium (methods 7 and 8).…”

Building native protein conformation from NMR backbone chemical shifts using Monte Carlo fragment assembly

“…The acquisition of RDCs requires comparatively little additional data collection time beyond backbone resonance assignments but provides considerable additional structural information (Prestegard et al 2000; Tolman et al 2001). Several structure determination methods based on RDC‐restrained models have been developed (Delaglio et al 2000; Andrec et al 2001; Rohl and Baker 2002; Kontaxis et al 2005; Mayer et al 2006). However, methods that rely primarily on RDC restraints require multiple data sets collected in different media to overcome restraint degeneracy.…”