title: Conformational analysis of g protein-coupled receptor signaling by hydrogen/deuterium exchange mass spectrometry.
abstract: Conformational change and protein-protein interactions are two major mechanisms of membrane protein signal transduction, including G protein-coupled receptors (GPCRs). Upon agonist binding, GPCRs change conformation, resulting in interaction with downstream signaling molecules such as G proteins. To understand the precise signaling mechanism, studies have investigated the structural mechanism of GPCR signaling using X-ray crystallography, nuclear magnetic resonance (NMR), or electron paramagnetic resonance. In addition to these techniques, hydrogen/deuterium exchange mass spectrometry (HDX-MS) has recently been used in GPCR studies. HDX-MS measures the rate at which peptide amide hydrogens exchange with deuterium in the solvent. Exposed or flexible regions have higher exchange rates and excluded or ordered regions have lower exchange rates. Therefore, HDX-MS is a useful tool for studying protein-protein interfaces and conformational changes after protein activation or protein-protein interactions. Although HDX-MS does not give high-resolution structures, it analyzes protein conformations that are difficult to study with X-ray crystallography or NMR. Furthermore, conformational information from HDX-MS can help in the crystallization of X-ray crystallography by suggesting highly flexible regions. Interactions between GPCRs and downstream signaling molecules are not easily analyzed by X-ray crystallography or NMR because of the large size of the GPCR-signaling molecule complexes, hydrophobicity, and flexibility of GPCRs. HDX-MS could be useful for analyzing the conformational mechanism of GPCR signaling. In this chapter, we discuss details of HDX-MS for analyzing GPCRs using the β2AR-G protein complex as a model system.
authors.affiliation: Department of Medicine, University of California at San Diego, San Diego, California, USA.
authors.forename: Sheng
authors.initials: S
authors.lastname: Li
authors.affiliation: School of Pharmacy, Sungkyunkwan University, Suwon, Republic of Korea.
authors.forename: Su Youn
authors.initials: SY
authors.lastname: Lee
authors.affiliation: School of Pharmacy, Sungkyunkwan University, Suwon, Republic of Korea. Electronic address: kychung2@skku.edu.
authors.forename: Ka Young
authors.initials: KY
authors.lastname: Chung
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D041961:Deuterium Exchange Measurement / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D025941:Protein Interaction Mapping / Q000379:methods*; D060066:Protein Interaction Maps; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D043562:Receptors, G-Protein-Coupled
keywords: GPCR; Hydrogen/deuterium exchange mass spectrometry; Protein conformation; Protein dynamics; Protein–protein interaction interface; Signaling proteins