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Solution NMR spectroscopy of GPCRs: Residue-specific labeling strategies with a focus on C-methyl methionine labeling of the atypical chemokine receptor ACKR3. Methods Cell Biol 2019;149:259-288

Date

01/09/2019

Pubmed ID

30616824

Pubmed Central ID

PMC6941889

DOI

10.1016/bs.mcb.2018.09.004

Scopus ID

2-s2.0-85056642454   1 Citation

Abstract

The past decade has witnessed remarkable progress in the determination of G protein-coupled receptor (GPCR) structures, profoundly expanding our understanding of how GPCRs recognize ligands, become activated, and interact with intracellular signaling components. In recent years, numerous studies have used solution nuclear magnetic resonance (NMR) spectroscopy to investigate GPCRs, providing fundamental insights into GPCR conformational changes, allostery, dynamics, and other facets of GPCR function are challenging to study using other structural techniques. Despite these advantages, NMR-based studies of GPCRs are few relative to the number of published structures, due in part to the challenges and limitations of NMR for the characterization of large membrane proteins. Several studies have circumvented these challenges using a variety of isotopic labeling strategies, including side chain derivatization and metabolic incorporation of NMR-active nuclei. In this chapter, we provide an overview of different isotopic labeling strategies and describe an in-depth protocol for the expression, purification, and NMR studies of the chemokine GPCR atypical chemokine receptor 3 (ACKR3) via CH-methionine incorporation. The goal of this chapter is to provide a resource to the GPCR community for those interested in pursuing NMR studies of GPCRs.

Author List

Kleist AB, Peterson F, Tyler RC, Gustavsson M, Handel TM, Volkman BF

Authors

Francis C. Peterson PhD Professor in the Biochemistry department at Medical College of Wisconsin
Brian F. Volkman PhD Professor in the Biochemistry department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Animals
Carbon Isotopes
Cell Line
Humans
Magnetic Resonance Spectroscopy
Receptors, CXCR
Receptors, G-Protein-Coupled
Solutions
Staining and Labeling
Vitamin U
jenkins-FCD Prod-444 eb4ebd1a08581aba961d3befd3b851a3c3ec6b46