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. 2022 Jul 29;13:4396. doi: 10.1038/s41467-022-31945-6

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© The Author(s) 2022

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — a Anisotropy decay curves of Alexa Fluor 555 (top row) at residue 175 (left) and 58 (right) and lifetime decay curves (bottom row) under magic angle conditions for apo (black) and holo state (green). b Same measurements as in (a) with Alexa Fluor 647 in apo (black) and holo state (red). c Anisotropy decay curves of TMR (top left) and Cy5 (top right) at residue 175 and lifetime decay curves of TMR (bottom left) and Cy5 (bottom right) under magic angle conditions for apo (black) and holo state (coloured). d FRET efficiency distributions (centre & bottom) of HiSiaP variant 175/228 for Alexa Fluor 555 – Alexa Fluor 647 (left) and TMR – Cy5 (right) in apo (grey) and holo state (green). e Converted distances from the mean FRET efficiencies are shown as black bars (mean of n = 3 independent experiments) in comparison to simulation (grey bar) and PELDOR/DEER results from Fig. 3. The red shade around the experimental smFRET distance are error bars based on the standard deviation of n = 3 independent experiments. Source data are provided as a Source Data File.