Determining the Role of Electrostatics in the Making and Breaking of the Caprin1-ATP Nanocondensate.

We employ a multiscale computational approach to investigate the condensation process of the C-terminal low-complexity region of the Caprin1 protein as a function of increasing ATP concentration for three states: the initial mixed state, nanocondensate formation, and dissolution of the droplet as it reenters the mixed state. We show that upon condensation, ATP assembles via pi-pi interactions, resulting in the formation of a large cluster of stacked ATP molecules stabilized by sodium counterions. The surface of the ATP assembly interacts with the arginine-rich regions of the Caprin1 protein, particularly with its N-terminus, to promote the complete phase-separated droplet on a length scale of tens of nanometers. In order to understand droplet stability, we analyzed the near-surface electrostatic potential (NS-ESP) of Caprin1 and estimated the zeta potential of the Caprin1-ATP assemblies. We predict a positive NS-ESP at the Caprin1 surface for low ATP concentrations that defines the early mixed state, in excellent agreement with the NS-ESP obtained from NMR experiments using paramagnetic resonance enhancement. By contrast, the NS-ESP of Caprin1 at the surface of the nanocondensate at moderate levels of ATP is highly negative compared to that at the mixed state, and estimates of a large zeta potential outside the highly dense region of charge further explain the remarkable stability of this phase-separated droplet assembly. As ATP concentrations rise further, the strong electrostatic forces needed for nanocondensate stability are replaced by weaker Caprin1-ATP interactions that drive the re-entry into the mixed state that exhibits a much lower zeta potential.