Department of Biochemistry and Molecular Biology, Monash University, Clayton Campus, Melbourne, Victoria, Australia.
PLoS One. 2013 Sep 30;8(9):e75835. doi: 10.1371/journal.pone.0075835. eCollection 2013.
We have developed an orange non-fluorescent photochromic protein (quantum yield, 0.003) we call Phanta that is useful as an acceptor in pcFRET applications. Phanta can be repeatedly inter-converted between the two absorbing states by alternate exposure to cyan and violet light. The absorption spectra of Phanta in one absorbing state shows excellent overlap with the emission spectra of a number of donor green fluorescent proteins including the commonly used EGFP. We show that the Phanta-EGFP FRET pair is suitable for monitoring the activation of caspase 3 in live cells using readily available instrumentation and a simple protocol that requires the acquisition of two donor emission images corresponding to Phanta in each of its photoswitched states. This the first report of a genetically encoded non-fluorescent acceptor for pcFRET.
我们开发了一种橙色非荧光光致变色蛋白（量子产率为 0.003），我们称之为 Phanta，它可用作 pcFRET 应用中的受体。Phanta 可以通过交替暴露于蓝光和紫光在两种吸收状态之间反复转换。Phanta 在一种吸收状态下的吸收光谱与包括常用的 EGFP 在内的许多供体绿色荧光蛋白的发射光谱具有极好的重叠。我们表明，Phanta-EGFP FRET 对适合使用现成的仪器和简单的协议监测活细胞中 caspase 3 的激活，该协议需要获取两个供体发射图像，对应于 Phanta 在其每个光开关状态下。这是第一个用于 pcFRET 的遗传编码非荧光受体的报告。
