Fig. 2: Biochemical and biophysical characterizations of XopR LLPS.

a Schematic diagram of the domains and charge pattern of XopR. IDR (upper panel) and charged residues (bottom panel) were analyzed by IUPRED2 and CIDER, respectively. b Representative SPR sensorgram of the XopR–XopR interaction. XopR at concentrations of 11 μM to 43 nM flowed over the chip with immobilized XopR. Binding parameters were generated using bivalent model. c Size exclusion chromatography of XopR at the indicated concentration of NaCl solution using Superdex 200 GL 10/300 Increase. The green dashed curve represents the elution profile of standard protein markers. d Liquid-liquid phase separation (LLPS) of XopR. XopR (10 μM, 10% XopR-mRuby2) was prepared in 50 mM NaCl solution, pH = 7.4, for 10 min before imaging. e XopR phase diagram was generated using ten images for each condition. Turbidity tests of XopR in solution are shown at the indicated NaCl concentration. f Typical force-distance profiles measured during approaching (open symbols) and separation (solid symbols) of the mica surfaces with an injected mixture of XopR coacervate as a function of the concentration of NaCl. g Representative SPR sensorgrams for XopR (on-chip) and AtFH1-FH1C, which were injected at concentrations of 20 μM to 39 nM. Binding parameters were generated using bivalent model. h Complex coacervation of XopR-AtFH1-FH1C in the low salt buffer (20 mM HEPES, 50 mM NaCl, pH = 7.4) and physiological buffer (20 mM HEPES, 150 mM NaCl, pH = 7.4). XopR (5 μM, 10% XopR-mRuby2) and AtFH1-FH1C (5 μM, 10% Alexa647-AtFH1-FH1C) were mixed for 10 min before imaging. i Effective interfacial energy of coacervates of XopR (10 μM), XopR-ScGFP (10 μM XopR + 10 μM ScGFP), and XopR-AtFH1-FH1C (10 μM XopR + 5 μM AtFH1-FH1C), as a function of NaCl concentration. Each of the effective surface energy values is averaged from three measurements.Scale bar: 10 μm in d, 1 µM for magnified images in d, and 10 μm in h.