Expression and purification of functional ligand-binding domains of T1R3 taste receptors

Chemical Senses
Yiling NieSteven D Munger

Abstract

Chemosensory receptors, including odor, taste, and vomeronasal receptors, comprise the largest group of G protein-coupled receptors (GPCRs) in the mammalian genome. However, little is known about the molecular determinants that are critical for the detection and discrimination of ligands by most of these receptors. This dearth of understanding is due in part to difficulties in preparing functional receptors suitable for biochemical and biophysical analyses. Here we describe in detail two strategies for the expression and purification of the ligand-binding domain of T1R taste receptors, which are constituents of the sweet and umami taste receptors. These class C GPCRs contain a large extracellular N-terminal domain (NTD) that is the site of interaction with most ligands and that is amenable to expression as a separate polypeptide in heterologous cells. The NTD of mouse T1R3 was expressed as two distinct fusion proteins in Escherichia coli and purified by column chromatography. Spectroscopic analysis of the purified NTD proteins shows them to be properly folded and capable of binding ligands. This methodology should not only facilitate the characterization of T1R ligand interactions but may also be useful for dissecting the funct...Continue Reading

Citations

Sep 18, 2012·Applied Microbiology and Biotechnology·Maud SigoillotLoïc Briand
May 1, 2007·Pflügers Archiv : European journal of physiology·Stephen D Roper
May 25, 2012·American Journal of Physiology. Regulatory, Integrative and Comparative Physiology·Yada Treesukosol, Alan C Spector
Dec 19, 2008·PloS One·Cedrick D DotsonSteven D Munger
Jan 28, 2014·PloS One·Becky R SimonOrmond A MacDougald
Jan 5, 2011·Nutrition Reviews·Eleanor R Grimm, Nanette I Steinle
Dec 23, 2008·Biochimica Et Biophysica Acta·Wei-Feng XueJannette Carey
Apr 1, 2008·Protein Expression and Purification·Yuxun Zhang, Xiang-Qun Xie
Feb 26, 2009·Journal of Food Science·B Meyers, M S Brewer
Mar 28, 2012·Protein Expression and Purification·Elodie MaîtrepierreLoïc Briand
Nov 3, 2016·Comparative Biochemistry and Physiology. Part D, Genomics & Proteomics·Sen GaoZhanjiang Liu
Sep 27, 2013·The Journal of Biological Chemistry·Becky R SimonOrmond A MacDougald
Feb 12, 2008·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Ken-ichiro NakajimaKeiko Abe
Jun 7, 2012·American Journal of Physiology. Endocrinology and Metabolism·Maartje C P GeraedtsSteven D Munger
Jun 7, 2021·Biochemical Pharmacology·Ksenia S KudryashovaAlexey V Feofanov

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