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. 2021 May 18;11:10536. doi: 10.1038/s41598-021-90039-3

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Distinct dimer conformations of WT and F220C opsins in the plasma membrane of HEK293 cells. (A) Fluorescence images of HEK293 cells expressing WT-FLAG-SNAP opsin and F220C-FLAG-SNAP opsin labeled with SNAP donor (BG-JF₅₄₉) and acceptor (BG-JF₆₄₆) dyes. The top and bottom rows show images of cells taken before and after bleaching of the acceptor by high intensity 640 nm illumination. Scale bar, 10 μm. (B) Summary of acceptor fluorescence as a reporter of comparable surface expression for both constructs. (C) Summary of donor fluorescence recovery upon acceptor bleaching, indicative of FRET (*, p = 0.007, unpaired t-test (two-tail)).