Technique Combo Beats NMR Solo

Researchers have combined nuclear magnetic resonance (NMR) with X-ray and neutron scattering to solve the largest protein solution structures to date. The structures illuminate how a key bacterial signal transduction pathway works and could lead to new antibiotics. The approach could also be useful for structural analysis of other large biomolecules.

Marius Clore of the National Institutes of Health and coworkers obtained the structures of the 128-kilodalton enzyme I (EI) dimer and its 146-kDa complex with histidine phosphocarrier protein (HPr) (J. Am. Chem. Soc., DOI: 10.1021/ja105485b). EI and HPr catalyze initial steps in a phosphotransfer process that transports sugar across bacterial cell membranes.

Biomolecular solution structures in the 50- to 80-kDa range have been solved by NMR alone, but the new study extends the accessible size range significantly by using multiple complementary solution techniques.

Crystal structures of EI had been solved previously, but they disagree with solution scattering data, indicating that domain orientations in crystals and in solution differ. The combined approach resolves these discrepancies and thus provides information that is not accessible from crystal structures alone.

In the combined technique, small- and wide-angle X-ray scattering provide shape and size information, NMR dipolar couplings (a type of internuclear interaction) indicate how to orient the domains properly, and small-angle neutron scattering validates the structures.

Other groups have used NMR and scattering techniques together, but Clore’s is the “first to demonstrate the combined approach experimentally on a biologically relevant high-molecular-weight complex,” says structural biologist Michael Sattler of the Technical University of Munich and Helmholtz Center Munich. And although complexes of similar size have been structurally analyzed, those studies were qualitative or focused on individual regions, whereas the Clore study “provides a full experimental description of a complete complex. It’s an impressive demonstration of the utility and success of hybrid approaches for structural analysis of challenging systems,” Sattler says.