Refinement of Multidomain Protein Structures by Combination of Solution Small-Angle X-ray Scattering and NMR Data

Abstract

Determination of the 3D structures of multidomain proteins by solution NMR methods presents a number of unique challenges related to their larger molecular size and the usual scarcity of constraints at the interdomain interface, often resulting in a decrease in structural accuracy. In this respect, experimental information from small-angle scattering of X-ray radiation in solution (SAXS) presents a suitable complement to the NMR data, as it provides an independent constraint on the overall molecular shape. A computational procedure is described that allows incorporation of such SAXS data into the mainstream high-resolution macromolecular structure refinement. The method is illustrated for a two-domain 177-amino-acid protein, γS crystallin, using an experimental SAXS data set fitted at resolutions from ∼200 Å to ∼30 Å. Inclusion of these data during structure refinement decreases the backbone coordinate root-mean-square difference between the derived model and the high-resolution crystal structure of a 54% homologous γB crystallin from 1.96 ± 0.07 Å to 1.31 ± 0.04 Å. Combining SAXS data with NMR restraints can be accomplished at a moderate computational expense and is expected to become useful for multidomain proteins, multimeric assemblies, and tight macromolecular complexes.