Salt-Dependent Rheology and Surface Tension of Protein Condensates Using Optical Traps

An increasing number of proteins with intrinsically disordered domains have been shown to phase separate in buffer to form liquidlike phases. These protein condensates serve as simple models for the investigation of the more complex membraneless organelles in cells. To understand the function of such proteins in cells, the material properties of the condensates they form are important. However, these material properties are not well understood. Here, we develop a novel method based on optical traps to study the frequency-dependent rheology and the surface tension of P-granule protein PGL-3 condensates as a function of salt concentration. We find that PGL-3 droplets are predominantly viscous but also exhibit elastic properties. As the salt concentration is reduced, their elastic modulus, viscosity, and surface tension increase. Our findings show that salt concentration has a strong influence on the rheology and dynamics of protein condensates suggesting an important role of electrostatic interactions for their material properties.

Figure 1

Microrheology setup based on a dual optical trap. (a) Schematic. (b) Fluorescent micrograph of protein droplet during the measurement. Scale bar, $5\mu \mathrm{m}$. (c) Mechanical circuit equivalent of measurement setup. (d) Exemplary experimental output during oscillatory droplet deformation: position of optical trap 1 (red) and forces associated to optical traps 1 and 2 (blue) at frequency 1 Hz and $75\mathrm{m}M$ KCl. (e) Exemplary system spring constants ${\chi }_{\text{sys}}^{\prime }$ (filled circles), ${\chi }_{\text{sys}}^{\prime \prime }$ (empty circles) from the measurement of a protein droplet with a diameter of $12.2\mu \mathrm{m}$ at a salt concentration of $75\mathrm{m}M$ KCl.

Figure 2

Spring constants for a Newtonian and non-Newtonian droplet subject to polar forcing. (a) Droplets are subject to an oscillatory pressure $P_0$ applied in a dome-shaped region at the poles. (b) Complex spring constant ${\chi }^{*}$ of a Newtonian droplet with surface tension. (c) Complex system spring constant of a Newtonian droplet in series with two identical traps. For low frequencies, the spring constant is dominated by surface tension (gray zone, left-hand side). For high frequencies, the two traps dominate the spring constant and ${\chi }_{\text{sys}}^{\prime }$ approaches $k/2$ (gray zone, right-hand side). Outside of this trap-dominated regime, the loss modulus is governed by the viscous properties of the droplet material. (d) Spring constant of a non-Newtonian droplet formed by a power law fluid, i.e., $G^{*}(\omega )=A\pi {\omega }^{\beta }[\csc (\pi \beta /2)+i\text{sec}(\pi \beta /2)]/2$, and with surface tension. Parameters are ${\theta }_0=0.1$, $R=5\mu \mathrm{m}$, $\gamma =3\mu \mathrm{N}/\mathrm{m}$, $\eta =0.5\mathrm{Pa}\mathrm{s}$, $A=0.2\mathrm{Pa}$, $\beta =0.8$, $k_1=k_2=150\mathrm{pN}/\mu \mathrm{m}$, and $\xi =6\pi {\eta }_{b}R$, where ${\eta }_b={10}^{-3}\mathrm{Pa}\mathrm{s}$.

Figure 3

Salt-dependent material properties of PGL-3 droplets. (a)–(b) Spring constants ${\chi }^{\prime }$ (filled circles), ${\chi }^{\prime \prime }$ (empty circles), and reconstructed moduli $G^{\prime }$ (filled circles), $G^{\prime \prime }$ (empty circles) for one droplet. [Corresponding ${\chi }_{\text{sys}}^{*}$ shown in Fig. 1] (c)–(f) Ensemble average of $G^{\prime }$ (closed symbols), $G^{\prime \prime }$ (open symbols) of the droplet material at salt concentrations of 75,115,150, and $180\mathrm{m}M$ KCl. Error bars indicate standard deviations ($n=3$). (g)–(j) Salt dependence of viscosity found using a linear fit to the loss modulus ($G^{\prime \prime }=\omega \eta$, where $\eta$ is the viscosity) (g), surface tension (h), storage modulus at 10 Hz (i), and loss tangent at 10 Hz (j). In panel (h), we show a fit of the theoretically predicted decay of surface tension in dependence of salt concentration $c$ (solid red line) [26], where $c_{cr}=243\mathrm{m}M$ is the fitted critical salt concentration.