Fig. 4: Time-dependent enzyme activity in cells with intracellular condensates.

a Schematic representation of the Renilla luciferase (Rluc)-catalyzed reaction, which oxidizes coelenterzine (CLZ) to coelenteramide (CLM) with concomitant emission of blue light. b Genetic constructs used for the expression of RIDD-tagged enzyme (Rluc-RIDD) and the phase- forming scaffold (FIB1-GFP-RIAD), or the RIDD-tagged enzyme in its free form (left). The engineered strain of Erluc1, containing intracellular condensates, is expected to exhibit an enhanced reaction rate shortly after condensate induction but a diminished rate over time compared to Erluc0 (right). c Comparison of Rluc catalytic efficiency between Erluc0 and Erluc1 at various point t_i following FIB1-GFP-RIAD induction. A.u. on the vertical axis refers to absorption units. d Comparison of relative reaction rates. The normalized reaction rates were calculated as the ratio of luminescence intensities between Erluc1 and Erluc0. The gray dashed line serves as a visual reference indicating that a relative reaction rate of 1 suggests comparable reaction rates between Erluc1 and Erluc0. e In-cell FRAP analysis of the intracellular condensates showing a gradual decrease in fluorescent recovery over time. E. coli cells expressing mCherry-ELK16 serve as a control for a solid matter. f FRAP recovery curves of intracellular condensates at different time points. Scale bar, 1 μm. g Immobile fractions of intracellular condensates normalized to mCherry-ELK16 aggregates, plotted as a function of time, derived from f. For c and d, data are represented as mean ± SD from three independent experiments (n = 3). For f and g, data are represented as mean ± SD from ten condensates (n = 10) from three independent experiments. Source data are provided as a Source Data file.