A General Mechanism of Photoconversion of Green-to-Red Fluorescent Proteins Based on Blue and Infrared Light Reduces Phototoxicity in Live-Cell Single-Molecule Imaging,Angewandte Chemie International Edition

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A General Mechanism of Photoconversion of Green-to-Red Fluorescent Proteins Based on Blue and Infrared Light Reduces Phototoxicity in Live-Cell Single-Molecule Imaging
Angewandte Chemie International Edition ( IF 16.1 ) Pub Date : 2017-07-17 06:26:06 , DOI: 10.1002/anie.201702870
Bartosz Turkowyd ₁ , Alexander Balinovic ₁ , David Virant ₁ , Haruko G. Gölz Carnero ₁ , Fabienne Caldana ₁ , Marc Endesfelder ₂ , Dominique Bourgeois ₃ , Ulrike Endesfelder ₁

Affiliation

Photoconversion of fluorescent proteins by blue and complementary near-infrared light, termed primed conversion (PC), is a mechanism recently discovered for Dendra2. We demonstrate that controlling the conformation of arginine at residue 66 by threonine at residue 69 of fluorescent proteins from Anthozoan families (Dendra2, mMaple, Eos, mKikGR, pcDronpa protein families) represents a general route to facilitate PC. Mutations of alanine 159 or serine 173, which are known to influence chromophore flexibility and allow for reversible photoswitching, prevent PC. In addition, we report enhanced photoconversion for pcDronpa variants with asparagine 116. We demonstrate live-cell single-molecule imaging with reduced phototoxicity using PC and record trajectories of RNA polymerase in Escherichia coli cells.

中文翻译：

基于蓝色和红外光的绿色到红色荧光蛋白的光转换的一般机制减少了活细胞单分子成像中的光毒性。

蓝光和互补的近红外光对荧光蛋白的光转换（称为引发转换（PC））是Dendra2最近发现的一种机制。我们证明，控制古细菌家族（Dendra2，mMaple，Eos，mKikGR，pcDronpa蛋白家族）的荧光蛋白的残基69处的苏氨酸在残基66处的精氨酸构象代表了促进PC的一般途径。已知会影响发色团柔韧性并允许可逆光开关的丙氨酸159或丝氨酸173突变会阻止PC。此外，我们报道了pcDronpa变体与天冬酰胺116的增强的光转换。我们证明了使用PC降低了光毒性的活细胞单分子成像，并记录了大肠杆菌细胞中RNA聚合酶的轨迹。

更新日期：2017-07-17

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