Figure 3. Decoupling the contribution of the dense phase chemical environment and the interfacial electric field to the catalytic function of condensates.

A, The hydrolysis reaction of p-nitrophenol phosphate (pNPP) results in the formation of p-nitrophenol (a yellow product) and phosphoric acid.

B, Schematic showing that regions of an IDP can have different intrinsic preferences for interacting localizing to the dense phase versus preferentially localizing to the interfaces of condensates.

C, Radial density plots of the N-terminal region (first four residues) and full-length protein from simulations of the full-length RLP_WT. Each data point represents mean ± SE. Inset shows the comparison of the density of the N-terminal sequence and the full-length sequence at the interface of the condensates based on the radial density plots. Bar graph shows the mean ± SE.

D, Comparison of the density of the N-terminal sequence at the condensate interface for different N-terminal sequences: Ser-Lys-Gly-Pro, Ser-Ser-Gly-Pro, and Ser-Tyr-Gly-Pro. Bar graph shows the mean ± SE.

E, Evaluation of the interfacial electric field of different sequences (SKGP-RLP, SSGP-RLP, SYGP-RLP) using ratiometric DI-4-ANEPPS fluorescence assay. The insets show the sample ratiometric images of condensates (at similar size) with DI-4-ANEPPS.

F, Fluorogenic hydrolysis reaction of resorufin phosphocholine generates fluorescent resorufin.

G, Comparison of the resorufin signal of the fluorogenic assay using different condensates formed by RLPs with different N-terminal sequences namely, SKGP-RLP, SSGP-RLP, and SYGP-RLP. The excitation was set at 550 nm with a WLL laser, and the emission detector was set at 570–600 nm on a HYD detector. The highest fluorescence signal within a 15 μm z range was identified for quantification of the resorufin signal. The grey line represents the spontaneous hydrolysis of resorufin phosphocholine in the same buffer solution. Scale bar is 5 μm. Each data point represents mean ± SE.