Fig. 3: LLPS condensate dynamics depend on the stability of CC building module interactions and can be modeled by a coarse-grained simulation.

a Designed sets of CC pairs with increased helical propensity comprising three or four heptads. b HEK293T cells expressing polypeptide pair of two types of CC-forming segments with three heptads, with gs linkers and CC segments arranged in a different pattern forming LLPS condensates. c Condensates exhibit fast fluorescence recovery in a FRAP experiment left yellow for Cit and right blue for BFP channel. Data are presented as mean ± SD (n = 9 for Cit and n = 7 for BFP, condensates from different cells examined over 3 independent experiments). Below is a picture of a bleached condensate at different time points. d Condensate fusion. Insets from Supplementary Movie 2. Merge of the cyan and yellow channel. e Snapshot at 20 μs from simulation of protein constructs in b, at 310 K, indicative of LLPS condensate. f HEK293T cells expressing a pair of two types of CC-forming segments composed of three heptads, with gs linkers arranged in the same sequential arrangement remain diffuse and do not form condensates. g Snapshot at 20 μs from simulation of protein constructs in f, at 310 K, indicating no LLPS. h Pair of two types of CC-forming segments composed of four heptads, with gs linkers and different arrangement of CC modules in each chain, formed condensates in HEK293T cells, (i) which are however largely immobile, with slow and low fluorescence recovery. Data are presented as mean ± SD left yellow graph for Cit and right blue for BFP channel (n  =  5 for Cit and n  =  3 for BFP, condensates from different cells examined over at least 2 independent experiments). Below, bleached condensate at different time points. j Snapshot at 20 μs from the simulation of 4-heptad protein combination in h, at 310 K, is indicative of LLPS condensate. k Mean square displacement (MSD) analysis of proteins with mixed coil types, comparing 3-heptad coil proteins versus 4-heptad coil proteins. MSD from 4-heptad coil proteins is additionally presented as an inset due to the difference of two orders of magnitude in MSD between the two types of proteins. Lines represent the mean values, and the shaded regions are the standard deviation (n = 3 simulation replicates). Scale bars, 10 μm. FRAP source data are provided as a Source Data file.