Amino-acid sequence of A1-LCD and molecular dynamics simulations of its condensation. (A) Amino-acid sequence. (B) First and (C, D) last frames from 1-μs simulations of the 8-chain systems at low and high salt. In all figures, Cl- or Na+ ions are represented by green and magenta spheres, respectively.

Salt effects on A1-LCD condensation and inter-chain interactions. (A) Dmax values, averaged over four replicates, as a function of simulation time. (B) Radii of gyration (Rg) from small-angle X-ray scattering10 and from MD simulations. Dashed curves are drawn to guide the eye. (C) Average number of inter- or intra-chain contacts per residue at each salt concentration. Dashed lines are drawn to show trends.

Levels of ion binding at different NaCl concentrations. (A) Number of 1st-shell ions, with (darker colors; tick marks on the left vertical axis) and without (lighter colors; tick marks on the right vertical axis) normalization by the number of residues of a given amino-acid type, at 1000 mM NaCl. (B) Total number of 1st-shell only or 1st- and 2nd-shell ions. (C) Net charge of the system with 1st- and 2nd-shell ions included.

Bridging ions. (A) Examples of chain bridging by Cl- and Na+; Cl- ions are coordinated to Arg and other sidechains whereas Na+ ions are coordinated to both backbone carbonyls (including from Gly) and sidechain oxygens. (B) Average number of Cl- or Na+ ions engaged in bridging between A1-LCD chains. (C) Average number of ions bound in 1st- and 2nd-shell sites lined by a given number of A1-LCD chains.

Indirect effects of ions on different types of interactions. (A) Number of inter-chain contacts per chain for each interaction type. Bars from left to right correspond to increasing salt concentrations (50, 150, 300, 500, and 1000 mM). Dashed lines are drawn to show trends. (B) An inter-chain salt bridge, with ion coordination by the partner sidechains. (C) An inter-chain π-π interaction, free of ion coordination. (D) Schematic showing a π-π interaction facilitated by high salt, via drawing water away from the interaction partners. (E) Radial distribution functions of water around Tyr residues that interact with Phe, Arg, Lys, Gln, and Asn. Lower values at high salt demonstrate water withdrawal.

Four classes of salt dependence and their prediction from amino-acid composition. (A) Salt dependences of liquid-liquid phase separation (LLPS). (B) Charge-charge and π-type interactions and their regulation by salt. (a) Significant Charge-charge attraction. (b) Screening of charge-charge attraction by salt. (c) Strengthening of π-type interactions by high salt. (d) Repulsion due to high net charge. (C) Distinctions of the four classes of salt dependence by three determinants: charged content (“Chg”), net charge (“Net”), and aromatic content (“Aro”).

Correlation between class of salt dependence and amino-acid composition