BRET evidence that β2 adrenergic receptors do not oligomerize in cells

Tien Hung Lan, Qiuju Liu, Chunman Li, Guangyu Wu, Jan Steyaert, Nevin A Lambert

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Bioluminescence resonance energy transfer (BRET) is often used to study association of membrane proteins, and in particular oligomerization of G protein-coupled receptors (GPCRs). Oligomerization of class A GPCRs is controversial, in part because the methods used to study this question are not completely understood. Here we reconsider oligomerization of the class A β2 adrenergic receptor (β2 AR), and reevaluate BRET titration as a method to study membrane protein association. Using inducible expression of the energy acceptor at multiple levels of donor expression we find that BRET between β2 AR protomers is directly proportional to the density of the acceptor up to ∼3,000 acceptors μm-2, and does not depend on the density of the donor or on the acceptor:donor (A:D) stoichiometry. In contrast, BRET between tightly-associating control proteins does not depend on the density of the acceptor, but does depend on the density of the donor and on the A:D ratio. We also find that the standard frameworks used to interpret BRET titration experiments rely on simplifying assumptions that are frequently invalid. These results suggest that β 2 ARs do not oligomerize in cells, and demonstrate a reliable method of assessing membrane protein association with BRET.

Original languageEnglish (US)
Article number10166
JournalScientific Reports
Volume5
DOIs
StatePublished - May 8 2015

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Energy Transfer
Adrenergic Receptors
Membrane Proteins
G-Protein-Coupled Receptors
Protein Subunits
varespladib methyl
Proteins

ASJC Scopus subject areas

  • General

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BRET evidence that β2 adrenergic receptors do not oligomerize in cells. / Lan, Tien Hung; Liu, Qiuju; Li, Chunman; Wu, Guangyu; Steyaert, Jan; Lambert, Nevin A.

In: Scientific Reports, Vol. 5, 10166, 08.05.2015.

Research output: Contribution to journalArticle

Lan, Tien Hung ; Liu, Qiuju ; Li, Chunman ; Wu, Guangyu ; Steyaert, Jan ; Lambert, Nevin A. / BRET evidence that β2 adrenergic receptors do not oligomerize in cells. In: Scientific Reports. 2015 ; Vol. 5.
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