Extended Data Fig. 4: Additional explorations on the nature of HS protein’s LLPS.
From: Artificial metalloenzyme assembly in cellular compartments for enhanced catalysis
a, Fluorescence recovery from the in vitro FRAP experiment indicating the liquid-like nature of the phase-separated HS protein. Additional in cellulo FRAP results and related discussions are provided in Supplementary Fig. 3 in the Supplementary Information. Scale bar = 50 µm. b, Confocal microscopy images of live compartmentalized HS–E. coli stained by Nile red (0.01% wt/vol) or thioflavin T (50 μM) for 1 h at 25 °C. The staining pattern suggested that the LLPS compartments should have provided a relatively hydrophobic microenvironment, which selectively uptakes the more hydrophobic dye, Nile red. n = 3 independent experiments; representative results are shown for each experiment described in a,b. Scale bar = 5 µm. c, LLPS compartment (labeled with Rho–BG) disruption by incubating with 1,6-hexanediol (1,6-HD, 20% wt/vol). 1,6-HD is known to partially dissolve phase-separated proteins by disrupting hydrophobic interactions within the droplets. This result suggested that hydrophobic interaction might be the primary driving force for the LLPS of HS protein. n = 4 biological replicates; representative images are shown. Scale bar = 5 μm. d, Statistics on 1,6-HD’s disruptive effect on LLPS compartments at different stages. n = 4 biological replicates. Data are presented as mean ± s.d.
