文献类型：期刊文献

英文题名：Sensitive detection via the time-resolved fluorescence of circularly permuted yellow fluorescent protein biosensors

作者：Li, Lei[1,3];Cheng, Yuchi[1];Shen, Shiyu[2,5];Zhou, Jiasheng[1];Wang, Aoxue[2,5];Chen, Guoqing[3];Xu, Jianhua[1];Yang, Yi[2,4];Zhao, Yuzheng[2,5];Zhang, Sanjun[1,6,7];Tian, Yang[8]

机构：[1]East China Normal Univ, State Key Lab Precis Spect, 3663 North Zhongshan Rd, Shanghai 200062, Peoples R China;[2]East China Univ Sci & Technol, Res Unit,Chinese Acad Med Sci, Optogenet & Synthet Biol Interdisciplinary Res Ct, Shanghai Collaborat Innovat Ctr Biomfg Technol,St, 130 Mei Long Rd, Shanghai 200237, Peoples R China;[3]Jiangnan Univ, Sch Sci, Wuxi 214122, Jiangsu, Peoples R China;[4]Chinese Acad Sci, CAS Ctr Excellence Brain Sci, Shanghai Inst Biol Sci, Shanghai 200031, Peoples R China;[5]East China Univ Sci & Technol, Sch Pharm, Shanghai Key Lab New Drug Design, 130 Mei Long Rd, Shanghai 200237, Peoples R China;[6]Shanxi Univ, Collaborat Innovat Ctr Extreme Opt, Taiyuan 030006, Shanxi, Peoples R China;[7]NYU Shanghai, NYU ECNU Inst Phys, 3663 North Zhongshan Rd, Shanghai 200062, Peoples R China;[8]East China Normal Univ, Sch Chem & Mol Engn, Dongchuan Rd 500, Shanghai 200241, Peoples R China

年份：2020

卷号：321

外文期刊名：SENSORS AND ACTUATORS B-CHEMICAL

收录：;EI(收录号：20203008978867);WOS:【SCI-EXPANDED(收录号:WOS:000562371800006)】；

基金：This work was supported by the National Natural Science Foundation of China (11774096, 11674101, 6141001145, 31722033, 31671484, 91649123, and 21827814), National Key Research and Development Program of China (2019YFA0904800, 2017YFA050400, 2018YFC1604204), Research Unit of New Techniques for Live-cell Metabolic Imaging (Chinese Academy of Medical Sciences, 2019RU01, 2019-I2M-5-013), Major Program of Development Fund for Shanghai Zhangjiang National Innovation Demonstration Zone (Stem Cell Strategic Biobank and Stem Cell Clinical Technology Transformation Platform, ZJ2018-ZD-004), Innovative research team of high-level local universities in Shanghai, the 111 project (B12024), and the Fundamental Research Funds for the Central Universities (JUSRP11837).

语种：英文

外文关键词：Circularly permuted fluorescent proteins; Time-resolved fluorescence; Biosensor; Fluorescent lifetime imaging

摘要：Circularly permuted fluorescent proteins (cpFP) have been used to develop diverse biosensors for monitoring cellular metabolites and states. In this article, we applied time-resolved spectroscopy to study a novel genetically encoded histidine biosensor, FHisJ, in vitro and in vivo. The average lifetime excited at 485 nm changes by 1.2 ns upon binding of histidine, which is much wider than most cpYFP-based biosensors. The ratio of fractional intensities changes 8-fold, which is higher than the excitation ratio measured at same conditions. More importantly, we study the photocycles of various cpYFP biosensors, i.e, FHisJ, SoNar, Frex and iNap. This work studies the difference of energy levels of cpYFP biosensors before and after binding with analytes, and reveals its effect on fluorescence intensity and lifetime. The cpYFP biosensors exist in 3 forms (A, I and B), but the fluorescence mainly comes from the excited states I* and B*. The dynamic range of the excitation ratio depends on the variation of A/(I+B) to some extent. However, the dynamic range of time-resolved fluorescence is related to the variation of I*/B*. This study will inspire the design of novel cpFP biosensors, especially those based on time resolved fluorescence.