Scattering of neutrons (basics)
EMBL Hamburg, Germany
Small angle neutron scattering (SANS) affords tremendous opportunities to extract structural parameters from a diverse range of macromolecules and macromolecular complexes in solution. The key to understanding SANS is the fundamental concept of contrast and how contrast can be systematically manipulated during the course of an experiment. Contrast, as the name implies, is simply the difference between a selected physical property of an object relative to the background in which the object is placed. For small angle X-ray scattering (SAXS), this selected property is is based in photon/electron interactions, i.e., X-ray scattering contrast is dependent on the difference between the electron density of a macromolecule and the electron density of the supporting solvent. For SANS, the scattering patterns are sourced from events between neutrons and the nuclei of atoms and thus the measured scattering intensities relate to the difference between the isotopic composition of a macromolecule and the isotopic composition of the solvent. Each isotope has a different probability to scatter an incident neutron. Thus, by varying the isotopic composition of either the macromolecule or the solvent, the contrast of a system can be increased or decreased. Importantly, the neutron scattering properties of the most common biological element, 1H, is very different to the other biological elements (12C, 14N, 16O, 31P and 32S) as well as the heavy hydrogen counterpart, deuterium, 2H. By changing the 1H per unit volume via 2H substitution (e.g., replacing 1H2O with 2H2O in the solvent) the contrast of a system can be altered and its effect on the scattering intensities evaluated.
The utility of SANS with contrast variation comes into play when investigating multi-component systems, for example higher-order macromolecular complexes and assemblies. If the individual components of a complex have different contrasts relative to the solvent and each other, it becomes possible to isolate the individual scattering contributions made by each component via contrast variation. From such contrast variation data, the shape and dispositions of the components within a complex can be determined while bound together. As a result, SANS has broad applications for the structural analysis of protein-RNA, protein-DNA and lipoprotein complexes as well as membrane systems, nanodiscs, metal-conjugated bionanoparticles, polymers and even more complicated systems that are otherwise inaccessible using X-rays. If biodeuteration has been employed to alter the 1H:2H ratio of a component, SANS with contrast variation can be used to determine the shapes of protein-protein, DNA-RNA complexes, etc. Here, the basics of neutron scattering are described along with practical aspects of sample preparation, including how to design an experiment, perform protein biodeuteration and what to do prior to and "at the beam line".
Texts and background
- Svergun, D.I., Koch, M.H., Timmins, P.A., & May, R. (2013) Small angle X-ray and neutron scattering from solutions of biological macromolecules. IUCr Texts on Crystallography, 19, Oxford Science Publications.
- Svergun, D.I., & Koch, M.H.J (2003) Small-angle scattering studies of biological macromolecules in solution. Rep. Prog. Phys. 66, 1735-1782.
- Whitten, A.E., Cai, S., and Trewhella, J. (2008) MULCh: ModULes for the analysis of small-angle neutron contrast variation data from biomolecular complexes. J. Appl. Cryst. 41, 222-226.
Protein-protein complexes and assemblies
- Whitten, A. E., Jacques, D. A., Hammouda, B., Hanley, T., King, G. F., Guss, J. M., Trewhella, J. and Langley, D. B. (2007) The structure of the sda-kinA complex suggests an allosteric mechanism of histidine kinase inhibition, J. Mol. Biol. 368, 407-420, 2007.
- Whitten, A.E., Jeffries, C.M., Harris, S.P., & Trewhella, J. (2008) Cardiac myosin binding protein-C decorates F-actin: Implications for cardiac function. Proc. Nat. Acad. Sci. USA. 105(47):18360-18365.
- To be discussed by Prof. Jill Trewhella – Joint Use of SAXS and SANS
- Capel M.S., Engelman D.M., Freeborn B.R., Kjeldgaard M., Langer J.A., Ramakrishnan V., Schindler D.G., Schneider D.K., Schoenborn B.P., Sillers I.Y., Yabuki, S & Moore, P.B.. (1987) A complete mapping of the proteins in the small ribosomal subunit of Escherichia coli. Science 238(4832):1403-6.
Lipids and nanodiscs
- Clifton L.A., Neylon C., & Lakey J.H. (2013) Examining protein-lipid complexes using neutron scattering. Methods Mol. Biol. 974:119-150.
- Frank Gabel, F., Lensink, M.F., Clantin, B., Jacob-Dubuisson, F., Villeret, V., & Ebel, C. (2014) Probing the conformation of FhaC with small-angle neutron scattering and molecular modeling. Biophys. J. 107(1): 185-196.
- Midtgaard S.R., Pedersen M.C., Kirkensgaard J.J., Sørensen K.K., Mortensen K., Jensen K.J., & Arleth L. (2014) Self-assembling peptides form nanodiscs that stabilize membrane proteins. Soft Matter. 10(5):738-52.
- Mehan S., Chinchalikar A.J., Kumar S., Aswal V.K, & Schweins R. (2013) Small-angle neutron scattering study of structure and interaction of nanoparticle, protein, and surfactant complexes. Langmuir. 29(36):11290-9.
- Goldenberg, D.P., & Argyle, B. (2014) Minimal effects of macromolecular crowding on an intrinsically disordered protein: A small-angle neutron scattering study. Biophys. J. 106(4), 905-914.
Date/time: Monday, 27 October 2014, 11:30