一种生物发光共振能量转移（BRET）系统：应用于相互作用的生物钟蛋白。
A bioluminescence resonance energy transfer (BRET) system: application to interacting circadian clock proteins.
作者信息
Xu Y, Piston D W, Johnson C H
机构信息
Department of Biology, Box 1812-B, Vanderbilt University, Nashville, TN 37235, USA.
出版信息
Proc Natl Acad Sci U S A. 1999 Jan 5;96(1):151-6. doi: 10.1073/pnas.96.1.151.
Abstract
We describe a method for assaying protein interactions that offers some attractive advantages over previous assays. This method, called bioluminescence resonance energy transfer (BRET), uses a bioluminescent luciferase that is genetically fused to one candidate protein, and a green fluorescent protein mutant fused to another protein of interest. Interactions between the two fusion proteins can bring the luciferase and green fluorescent protein close enough for resonance energy transfer to occur, thus changing the color of the bioluminescent emission. By using proteins encoded by circadian (daily) clock genes from cyanobacteria, we use the BRET technique to demonstrate that the clock protein KaiB interacts to form homodimers. BRET should be particularly useful for testing protein interactions within native cells, especially with integral membrane proteins or proteins targeted to specific organelles.
摘要
我们描述了一种检测蛋白质相互作用的方法，该方法相较于以往的检测方法具有一些吸引人的优势。这种方法称为生物发光共振能量转移（BRET），它使用一种与一种候选蛋白基因融合的生物发光荧光素酶，以及一种与另一种感兴趣的蛋白融合的绿色荧光蛋白突变体。两种融合蛋白之间的相互作用可使荧光素酶和绿色荧光蛋白足够靠近，从而发生共振能量转移，进而改变生物发光发射的颜色。通过使用来自蓝细菌的昼夜节律（每日）时钟基因编码的蛋白质，我们利用BRET技术证明时钟蛋白KaiB相互作用形成同源二聚体。BRET对于检测天然细胞内的蛋白质相互作用应该特别有用，尤其是对于整合膜蛋白或靶向特定细胞器的蛋白质。
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