Revealing the Hidden Sensitivity of Intrinsically Disordered Proteins to their Chemical Environment

Abstract

Intrinsically disordered protein-regions (IDRs) make up roughly 30% of the human proteome and are central to a wide range of biological processes. Given a lack of persistent tertiary structure, all residues in IDRs are, to some extent, solvent exposed. This extensive surface area, coupled with the absence of strong intramolecular contacts, makes IDRs inherently sensitive to their chemical environment. We report a combined experimental, computational, and analytical framework for high-throughput characterization of IDR sensitivity. Our framework reveals that IDRs can expand or compact in response to changes in their solution environment. Importantly, the direction and magnitude of conformational change depend on both protein sequence and cosolute identity. For example, some solutes such as short polyethylene glycol chains exert an expanding effect on some IDRs and a compacting effect on others. Despite this complex behavior, we can rationally interpret IDR responsiveness to solution composition changes using relatively simple polymer models. Our results imply that solution-responsive IDRs are ubiquitous and can provide an additional layer of regulation to biological systems.

Figure 1.

(A) Fluorescence spectra normalized to donor peak intensity of a FRET construct in compacting (red), buffer (black), and expanding (blue) solutions. Cyan and green areas are base spectra of donor and acceptor FPs, respectively. Inset shows single configurations for various degrees of expansion. (B) FRET efficiency of Gly-Ser repeat linkers vs. number of residues (N) in a buffer solution. UT is a solution of untethered, equimolar donor and acceptor. Dashed line shows linear fit of the data. (C) Calculated χ for FRET constructs in buffer determined by experiment (average of four repeats with 6 replicates each) and simulation (average of five repeats). Error bars are SD of all replicates/repeats.

Figure 2.

(A) Solution space scans of IDRs. Each data point shows the average χ vs. concentration of a specific solute for each protein taken from two repeats. Vertical grey bars show spread of data, and are often too small to see. Proteins vary down columns, and solutes across rows. Background color represents the sensitivity of change to solute addition: stronger colors imply higher sensitivity, red hues indicate compaction, and blue hues indicate expansion. Purple background indicates non-monotonic behavior. (B) Differential response of IDRs to individual solutes. Each panel point shows Δχ = χ([solute]) − χ([solute] = 0) vs. concentration from two repeats of a specific solute for several different constructs. Vertical lines are the spread of the data.

Figure 3.

All-atom simulations of IDR sensitivity to solutions. (A) Heatmap of protein sensitivity and molecular features for all 70 IDR sequences simulated. Protein identity varies from top to bottom across cells, and molecular features vary left to right. Color-maps are shown for each molecular feature. (B) The magnitude Δχ in attractive (blue) or repulsive (red) solutions as a function of χ in aqueous solution for each protein in (A). Error bars calculated from SD of 5 repeats. All data available in Table S3.

Fig. 4

(A–C). Density maps of all-atom simulations shown in Fig, 3B (A), PIMMS coarse-grained simulations (B), and an analytical model (C) for solution sensitivity Δχ vs dimensions in aqueous buffer χ. (D) Coil-to-globule transition obtained from an analytical model (SARC = self-avoiding random coil). Δχ is measured as the height of the blue (contraction) or red (expansion) shaded regions. When the same chain-solvent perturbation (Δ_sol) is applied to a 100-residue chain with different starting χ values, very different Δχ are expected. (E) Projection of experimental data for Ash1 onto the analytical model from (D), with solute concentrations scaled to the change in mean-field chain-solvent interaction as compared with neat buffer. The x-axis here represents the same units as in panel (D) but is offset so that chain:solvent interactions in aqueous solvent are set to 0. Chain dimensions are also shown by their apparent scaling exponent ν^app. Mapping of other proteins is shown in Fig. S7.