Kliegman, R. | bioRxiv Science

bioRxiv2026

The exchange dynamics of client molecules in biomolecular condensates

Biomolecular condensates are dynamic assemblies whose functions depend on continuous exchange of molecular components with the surrounding environment. While scaffold molecules drive phase separation and condensate architecture, many functional components are clients that are recruited through interactions with the scaffold-rich environment. Despite their prevalence, how client- scaffold interactions shape client exchange dynamics remains poorly understood. Here, we develop a reaction-diffusion model for client exchange in scaffold-driven condensates, in which clients switch between a scaffold-bound state and an unbound state. Bound clients exchange through scaffold-mediated transport, whereas unbound clients diffuse through the pore space of the condensate. Using the fluorescence recovery of fully photobleached condensates as a measure of client exchange, we compare transport through these two pathways with bound-unbound conversion and identify three limiting regimes. In the slow-conversion regime, bound and unbound clients recover through distinct scaffold- and pore-mediated pathways. In the intermediate-conversion regime, recovery of bound clients becomes limited by client unbinding. In the fast-conversion regime, local equilibrium between bound and unbound clients produces an effective single-state recovery. We further propose a unifying description that connects these regimes and quantitatively captures the apparent recovery timescales extracted from numerical simulations across condensate sizes. Our results provide a framework for interpreting component-specific exchange dynamics, and highlight client size, client- scaffold binding, and condensate porosity as key regulators of client turnover in multicomponent condensates.

biophysics↗

bioRxiv2025

Dissecting Rate-Limiting Processes in Biomolecular Condensate Exchange Dynamics

An increasing number of biomolecules have been shown to phase-separate into biomolecular condensates -- membraneless subcellular compartments capable of regulating distinct biochemical processes within living cells. The speed with which they exchange components with the cellular environment can influence how fast biochemical reactions occur inside condensates and how fast condensates respond to environmental changes, thereby directly impacting condensate function. While Fluorescence Recovery After Photobleaching (FRAP) experiments are routinely performed to measure this exchange timescale, it remains a challenge to distinguish the various physical processes limiting fluorescence recovery and identify each associated timescale. Here, we present a reaction-diffusion model for condensate exchange dynamics and show that such exchange can differ significantly from that of conventional liquid droplets due to the presence of a percolated molecular network, which gives rise to different mobility species in the dense phase. In this model, exchange can be limited by diffusion of either the high- or low-mobility species in the dense phase, diffusion in the dilute phase, or the attachment/detachment of molecules to/from the network at the surface or throughout the bulk of the condensate. Through a combination of analytic derivations and numerical simulations, we quantify the contributions of these distinct physical processes to the overall exchange timescale and predict an experimentally testable scaling relationship between the exchange timescale and condensate size. We discover that the exchange dynamics can be accelerated via a pore-mediated pathway in which molecules pass through the pores of the meshwork and attach/detach directly in the condensate interior. Notably, this pathway leads to a new regime in which the exchange timescale becomes independent of condensate size, which we validate through FRAP experiments on a biosynthetic DNA nanostar system. Our work offers insight into the predominant physical mechanisms driving condensate material exchange, with implications for natural and engineered systems.

biophysics↗