文章基本信息

标题：BRET evidence that β2 adrenergic receptors do not oligomerize in cells
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作者：Tien-Hung Lan ; Qiuju Liu ; Chunman Li 等
期刊名称：Scientific Reports
电子版ISSN：2045-2322
出版年度：2015
卷号：5
DOI：10.1038/srep10166
出版社：Springer Nature
摘要：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 (β₂AR), 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 β₂AR 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 β₂ARs do not oligomerize in cells, and demonstrate a reliable method of assessing membrane protein association with BRET.