Mechanism of voltage-sensitive fluorescence in a microbial rhodopsin

Maclaurin D.; Venkatachalam V.; Lee H.Cohen A.E.

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Mechanism of voltage-sensitive fluorescence in a microbial rhodopsin

机译：Mechanism of voltage-sensitive fluorescence in a microbial rhodopsin

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Microbial rhodopsins were recently introduced as genetically encoded fluorescent indicators of membrane voltage. An understanding of the mechanism underlying this function would aid in the design of improved voltage indicators. We asked, what states can the protein adopt, and which states are fluorescent? How does membrane voltage affect the photostationary distribution of states? Here, we present a detailed spectroscopic characterization of Archaerhodopsin 3 (Arch). We performed fluorescence spectroscopy on Arch and its photogenerated intermediates in Escherichia coli and in single HEK293 cells under voltage-clamp conditions. These experiments probed the effects of time-dependent illumination and membrane voltage on absorption, fluorescence, membrane current, and membrane capacitance. The fluorescence of Arch arises through a sequential three-photon process. Membrane voltage modulates protonation of the Schiff base in a 13-cis photocycle intermediate (M ? N equilibrium), not in the ground state as previously hypothesized. We present experimental protocols for optimized voltage imaging with Arch, and we discuss strategies for engineering improved rhodopsin-based voltage indicators.

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《Proceedings of the National Academy of Sciences of the United States of America》 |2013年第15期|5939-5944|共6页
作者
Maclaurin D.; Venkatachalam V.; Lee H.Cohen A.E.;
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作者单位

Department of Physics, Harvard University, Cambridge, MA 02138, United States;

Graduate Program in Biophysics, Harvard University, Cambridge, MA 02138, United States;

Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138, United States;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
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正文语种英语
中图分类自然科学总论;
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Mechanism of voltage-sensitive fluorescence in a microbial rhodopsin

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