Nondenaturing mass spectrometry to study noncovalent protein/protein and protein/ligand complexes: technical aspects and application to the determination of binding stoichiometries.
In the present chapter we detail how mass spectrometry (MS) can be used to characterize noncovalent complexes, especially multimeric proteins and protein/ligand complexes. This original application of MS, also called "supramolecular MS" or "nondenaturing MS," appeared in the early 1990s and has continuously evolved since then. Nondenaturing MS is now fully integrated in structural biology programs and in drug discovery platforms. Indeed, appropriate sample preparation and fine tuning of the instrument make it possible to transfer weak assemblies without disruption from solution into the gas phase of the mass spectrometer. In this chapter we detail experimental conditions (sample preparation, optimization of instrumental parameters, etc.) required for the detection of noncovalent complexes by MS. We then focus on the type of information and accuracy that we get after interpreting electrospray ionization mass spectra obtained under nondenaturing conditions, with emphasis on the determination of the stoichiometry of protein/protein and protein/ligand complexes.
Sanglier
Sarah
S
Laboratoire de Spectrométrie de Masse Bio-Organique, Institut Pluridisciplinaire Hubert Curien UMR 7178 CNRS, UniversitéLouis Pasteur, Strasbourg, France.
Atmanene
Cédric
C
Chevreux
Guillaume
G
Dorsselaer
Alain Van
AV
D000449:Aldehyde Reductase / Q000037:antagonists & inhibitors / Q000737:chemistry; D001426:Bacterial Proteins / Q000032:analysis; D008024:Ligands; D013058:Mass Spectrometry / Q000379:methods*; D046912:Multiprotein Complexes / Q000737:chemistry*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000032:analysis; D015203:Reproducibility of Results
D001426:Bacterial Proteins; D008024:Ligands; D046912:Multiprotein Complexes; D000449:Aldehyde Reductase; C110847:HPr kinase; D017346:Protein Serine-Threonine Kinases