Sequence determinants of protein phase behavior from a coarse-grained model

Abstract

Membraneless organelles important to intracellular compartmentalization have recently been shown to comprise assemblies of proteins which undergo liquid-liquid phase separation (LLPS). However, many proteins involved in this phase separation are at least partially disordered. The molecular mechanism and the sequence determinants of this process are challenging to determine experimentally owing to the disordered nature of the assemblies, motivating the use of theoretical and simulation methods. This work advances a computational framework for conducting simulations of LLPS with residue-level detail, and allows for the determination of phase diagrams and coexistence densities of proteins in the two phases. The model includes a short-range contact potential as well as a simplified treatment of electrostatic energy. Interaction parameters are optimized against experimentally determined radius of gyration data for multiple unfolded or intrinsically disordered proteins (IDPs). These models are applied to two systems which undergo LLPS: the low complexity domain of the RNA-binding protein FUS and the DEAD-box helicase protein LAF-1. We develop a novel simulation method to determine thermodynamic phase diagrams as a function of the total protein concentration and temperature. We show that the model is capable of capturing qualitative changes in the phase diagram due to phosphomimetic mutations of FUS and to the presence or absence of the large folded domain in LAF-1. We also explore the effects of chain-length, or multivalency, on the phase diagram, and obtain results consistent with Flory-Huggins theory for polymers. Most importantly, the methodology presented here is flexible so that it can be easily extended to other pair potentials, be used with other enhanced sampling methods, and may incorporate additional features for biological systems of interest. 1/24. CC-BY-NC-ND 4.0 International license peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/238170 doi: bioRxiv preprint first posted online Dec. 21, 2017; Author summaryLiquid liquid phase separation (LLPS) of low-complexity protein sequences has emerged as an important research topic due to its relevance to membraneless organelles and intracellular compartmentalization. However a molecular level understanding of LLPS cannot be easily obtained by experimental methods due to difficulty of determining structural properties of phase separated protein assemblies, and of choosing appropriate mutations. Here we advance a coarse-grained computational framework for accessing the long time scale phase separation process and for obtaining molecular details of LLPS, in conjunction with state of the art enhanced sampling methods. We are able to capture qualitatively the changes of phase diagram due to specific mutations, inclusion of a folded domain, and to variation of chain length. The model is flexible and can be used with different knowledge-based potential energy func...

TDP-43 α-helical structure tunes liquid-liquid phase separation and function

“…To further elucidate the potential contributions of hydrophobicity vs. helicity to TDP-43 phase separation, we conducted coarse-grained simulations of TDP-43 CTD coexistence following the simulation model and framework from our previous work [46][47][48] . Because this coarse-grained model treats protein chains as flexible polymers and secondary structure is not directly captured, this allows us to decouple the mutational change in hydrophobicity from the changes in helicity, which are inseparable in experiment and traditional all-atom molecular simulation.…”

“…Coarse-grained simulations were conducted using an amino-acid-resolution model with 20 residue types to capture sequence specificity, having interactions based on relative hydropathies of each amino acid 48,70 . Each system was simulated at a range of temperatures using constant volume and temperature using a Langevin thermostat, and slab geometry, which allows for efficient sampling of coexistence densities.…”

Tyrosine phosphorylation regulates hnRNPA2 granule protein partitioning & reduces neurodegeneration

“…Local translation is important in many cell types but is critical for myelination by oligodendrocytes and neuronal functions, including synaptic plasticity and axon guidance. Different kinds of mRNA transport granules likely exist, with distinct mRNA cargos, including neuronal β-actin mRNA transport granules containing zipcode binding protein/IGF2BP1 (Elvira et al, 2006;Kiebler and Bassell, 2006;Zhang et al, 2001) and hnRNPA2-containing granules transporting myelin basic protein mRNA in oligodendrocytes or Arc, neurogranin, and αCamKII in neurons (Ainger et al, 1993;Brumwell et al, 2002;Gao et al, 2008;Shan et al, 2003). Several other protein components of hnRNPA2containing transport granules have been identified, including hnRNPF, hnRNPAB, hnRNPE1, hnRNPK, and the microtubule associated protein ch-TOG (CKAP5) (Francone et al, 2007;Kosturko et al, 2005;Kosturko et al, 2006;Laursen et al, 2011;Raju et al, 2011;Torvund-Jensen et al, 2014;White et al, 2012).…”

Structure of biomolecular condensates from dissipative particle dynamics simulations

“…31 In an effort to go beyond this limitation, coarse-grained simulation techniques have been used in which the IDPs are simplified to linear chains of beads connected by springs. [32][33][34] Reversible binding sites are represented as a direct attraction between selected beads or as an effective hydrophobic repulsion from the solvent. When the attractive beads are placed at the ends of the molecules they form so-called telechelic polymers.…”