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Ligand-induced conformational changes in a SMALP-encapsulated GPCR

Biochim Biophys Acta Biomembr. 2020; 
Routledge SJ, Jamshad M, Little HA, Lin YP, Simms J, Thakker A, Spickett CM, Bill RM, Dafforn TR, Poyner DR, Wheatley M.
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Molecular Biology Reagents … 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5yl amino]ethyl)phenol} was purchased from Tocris, The mutant constructs [W246Y]A 2A R and [W268Y]A 2A R were custom synthesised and cloned into the EcoRI-NotI sites of the vector pPICZαA by Genscript 22 … Get A Quote

摘要

The adenosine 2A receptor (A2AR), a G-protein-coupled receptor (GPCR), was solubilised and purified encapsulated in styrene maleic acid lipid particles (SMALPs). The purified A2AR-SMALP was associated with phospholipids characteristic of the plasma membrane of Pichia pastoris, the host used for its expression, confirming that the A2AR-SMALP encapsulated native lipids. The fluorescence spectrum of the A2AR-SMALP showed a characteristic broad emission peak at 330 nm, produced by endogenous Trp residues. The inverse agonist ZM241385 caused 30% increase in fluorescence emission, unusually accompanied by a red-shift in the emission wavelength. The emission spectrum also showed sub-peaks at 321 nm, 335 nm and 3... More

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