Chapter 1

Atomic Structural Investigations of Self-Assembled Protein Complexes by Solid-State NMR

Antoine Loquet, James Tolchard and Birgit Habenstein

Abstract

Protein self-assemblies play essential roles in many biological processes ranging from bacterial and viral infections to basic cellular functions. They can be found in a wide range of supramolecular architectures, in homomeric and heteromeric forms, and often in symmetric arrangements. The biological function will then be dictated by the structure of the assembled object rather than by the subunits. Atomicresolution structural investigations of protein assemblies can be tedious because of their size, their insolubility, and often their non-crystallinity. Solid-state NMR (ssNMR) is a powerful technique used to obtain high-resolution structural models of these complex assemblies and to study their assembly processes and interactions at the atomic level. Unrestricted by object size or solubility, ssNMR can be applied to study the structures and interactions of macromolecular assemblies such as proteins in a membrane environment, protein filaments, pores, fibrils or oligomeric species. This chapter focusses on the established methods and recent advances in magic angle spinning (MAS) ssNMR for the detection of structural restraints in macromolecular protein assemblies and the determination of their atomic-resolution models. We will review different 13C and 15N isotope labelling approaches necessary to detect and differentiate intra- and intermolecular distance restraints that define the protein subunit structure and their relevance in the context of symmetric assemblies. The collection, and interpretation of, structural restraints in protein assemblies by ssNMR will be discussed. We will also introduce the recent developments in ultra-fast MAS ssNMR to study and determine atomic structures of sub-milligram quantities of molecular assemblies using proton detection. Finally, our aim is to also illustrate the complementarity of ssNMR to other techniques in structural biology such as solutionstate NMR, mass-per-length scanning transmission electron microscopy (STEM) measurements and cryo-electron microscopy.

Total Pages: 1-39 (39)

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