English
 
User Manual Privacy Policy Disclaimer Contact us
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT

Released

Journal Article

Structural investigations of molecular machines by solid-state NMR.

MPS-Authors
/persons/resource/persons36494

Loquet,  A.
Research Group of Solid-State NMR, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons79129

Habenstein,  B.
Research Group of Solid-State NMR, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons15414

Lange,  A.
Department of NMR Based Structural Biology, MPI for Biophysical Chemistry, Max Planck Society;

External Ressource
Fulltext (public)
There are no public fulltexts stored in PuRe
Supplementary Material (public)
There is no public supplementary material available
Citation

Loquet, A., Habenstein, B., & Lange, A. (2013). Structural investigations of molecular machines by solid-state NMR. Accounts of Chemical Research, 46(9), 2070-2079. doi:10.1021/ar300320p.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0014-5EC9-B
Abstract
Essential biological processes such as cell motion, signaling,protein synthesis, and pathogen-host interactions rely on multifunctional molecular machines containing supramolecular assemblies, that is, noncovalently assembled protein subunits. Scientists would like to acquire a detailed atomic view of the complete molecular machine to understand its assembly process and functions. Structural biologists have used various approaches to obtain structural information such as X-ray crystallography, solution NMR, and electron microscopy. The inherent insolubility and large size of these multicomponent assemblies restrict the use of solution NMR, and their noncrystallinity and elongated shapes present obstacles to X-ray crystallography studies. Not limited by molecular weight or crystallinity, solid-state NMR (ssNMR) allows for structural investigations of supramolecular assemblies such as helical filaments, cross-β fibrils, or membrane-embedded oligomeric proteins. In this Account, we describe recent progress in the application of ssNMR to the elucidation of atomic structures of supramolecular assemblies. We highlight ssNMR methods to determine the subunit interfaces in symmetric arrangements. Our use of [1-(13)C]- or [2-(13)C]-glucose as a carbon source during bacterial protein expression results in significant (13)C spin dilution that drastically improves the spectral quality and enables us to detect meaningful structural restraints. Moreover, we can unequivocally determine intermolecular restraints using mixed [(1:1)1-(13)C/2-(13)C]-glucose labeled assemblies. We recently illustrated the power of this methodology with the structure determination of the type III secretion system (T3SS) needle. One crucial aspect in elucidating the atomic structure of these large multicomponent complexes is to determine the subunit-subunit interfaces. Notably, we could probe the needle subunit interfaces by collecting (13)C-(13)C intermolecular restraints. In contrast, these interfaces are not accessible via high-resolution cryo-EM. This approach is readily applicable to other supramolecular assemblies containing symmetrically repeating protein subunits, and could be combined with other techniques to get a more complete picture of multicomponent structures. To determine near-atomic structures of assemblies of biological interest, researchers could combine ssNMR data collected at the subunit interfaces with the envelope obtained from cryo-EM and potentially with monomeric subunit crystal structures.