Introducing SEC-SANS for studies of complex self-organized biological systems.

Abstract

Small-angle neutron scattering (SANS) is maturing as a method for studying complex biological structures. Owing to the intrinsic ability of the technique to discern between 1H- and 2H-labelled particles, it is especially useful for contrast-variation studies of biological systems containing multiple components. SANS is complementary to small-angle X-ray scattering (SAXS), in which similar contrast variation is not easily performed but in which data with superior counting statistics are more easily obtained. Obtaining small-angle scattering (SAS) data on monodisperse complex biological structures is often challenging owing to sample degradation and/or aggregation. This problem is enhanced in the D2O-based buffers that are typically used in SANS. In SAXS, such problems are solved using an online size-exclusion chromatography (SEC) setup. In the present work, the feasibility of SEC-SANS was investigated using a series of complex and difficult samples of membrane proteins embedded in nanodisc particles that consist of both phospholipid and protein components. It is demonstrated that SEC-SANS provides data of sufficient signal-to-noise ratio for these systems, while at the same time circumventing aggregation. By combining SEC-SANS and SEC-SAXS data, an optimized basis for refining structural models of the investigated structures is obtained.