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. 2023 Sep 6;14:5444. doi: 10.1038/s41467-023-41241-6

Fig. 5. FRET measurements of AF350/AF488-labeled tau variants reveal conformational changes in tau upon MB-induced LLPS.

Fig. 5

a Schematic illustration of the fluorescence labeling sites in the four tau variants. b FRET spectra of tau variants in 50 mM Tris (pH 7.4). Black: 10 μM tau (no phase separation); green: 10 μM tau + 100 μM MB (no phase separation); blue: 10 μM tau + 5% PEG8000 (phase separation); red: 10 μM tau + 100 μM MB + 5% PEG8000 (strong phase separation). c Two-photon microscopy images of AF350/AF488-labeled tau17/244 with or without 100 μM MB in 50 mM Tris (pH 7.4) with 5% PEG8000. Scale bars, 10 μm. d Apparent FRET efficiency of tau droplets measured by the two-photon microscopy images under excitation of AF350 by 700 nm laser. Individual data points represent each droplet. A total of 58 droplets were analyzed. The bars indicate minimum and maximum. The center and bounds of box indicate median and SD, respectively. Significance levels were determined by unpaired two-sided Student’s t test. e The FRET spectra of tau variants in 20 mM HEPES (pH 7.4) without PEG8000. Black: 15 μM tau (weak phase separation); blue: 15 μM tau + 20 μM MB (weak phase separation); red: 15 μM tau and 100 μM MB (strong phase separation). A concentration of 0.2 μM AF350/AF488-labeled tau variants was mixed as dopant in all the above solutions for fluorescence spectra measurements and imaging. Source data are provided as a Source data file.