bims-indpro Biomed News
on Intrinsically disordered proteins
Issue of 2022‒04‒10
thirteen papers selected by
Sara Mingu
Johannes Gutenberg University
  1. RNA chain length and stoichiometry govern surface tension and stability of protein-RNA condensates.
  2. Global Structure of the Intrinsically Disordered Protein Tau Emerges from Its Local Structure.
  3. Improving Martini 3 for Disordered and Multidomain Proteins.
  4. Long-range structural preformation in yes-associated protein precedes encounter complex formation with TEAD.
  5. Dynamic Heterogeneity at the Interface of an Intrinsically Disordered Peptide.
  6. The small aromatic compound SynuClean-D inhibits the aggregation and seeded polymerization of multiple α-synuclein strains.
  7. In vitro characterization and molecular dynamics simulation reveal mechanism of 14-3-3ζ regulated phase separation of the tau protein.
  8. Implementation of residue-level coarse-grained models in GENESIS for large-scale molecular dynamics simulations.
  9. Emerging Roles for Phase Separation of RNA-Binding Proteins in Cellular Pathology of ALS.
  10. Resources for computational prediction of intrinsic disorder in proteins.
  11. PLETHORA-WOX5 interaction and subnuclear localization control Arabidopsis root stem cell maintenance.
  12. Functional inhibition of c-Myc using novel inhibitors identified through "hot spot" targeting.
  13. A histidine cluster determines YY1-compartmentalized coactivators and chromatin elements in phase-separated enhancer clusters.
  1. iScience. 2022 Apr 15. 25(4): 104105
    RNA chain length and stoichiometry govern surface tension and stability of protein-RNA condensates.
    Rabia Laghmach, Ibraheem Alshareedah, Matthew Pham, Muralikrishna Raju, Priya R Banerjee, Davit A Potoyan.
      Proteomic studies have shown that cellular condensates are frequently enriched in diverse RNA molecules, which is suggestive of mechanistic links between phase separation and transcriptional activities. Here, we report a systematic experimental and computational study of thermodynamic landscapes and interfacial properties of protein-RNA condensates. We have studied the affinity of protein-RNA condensation as a function of variable RNA sequence length and RNA-protein stoichiometry under different ionic environments and external crowding. We have chosen the PolyU sequences for RNA and arginine/glycine-rich intrinsically disordered peptide (RGG) for proteins as a model system of RNA-protein condensates, which we then investigate through in vitro microscopy measurements and coarse-grained molecular dynamics simulations. We find that crowding and RNA chain length can have a major stabilizing effect on the condensation. We also find that the RNA-protein charge ratio is a crucial variable controlling stability, interfacial properties, and the reentrant phase behavior of RGG-RNA mixtures.
    Keywords:  Biophysical Chemistry; Biophysical chemistry; Biophysics; Physical chemistry
    DOI:  https://doi.org/10.1016/j.isci.2022.104105
  2. JACS Au. 2022 Mar 28. 2(3): 673-686
    Global Structure of the Intrinsically Disordered Protein Tau Emerges from Its Local Structure.
    Lukas S Stelzl, Lisa M Pietrek, Andrea Holla, Javier Oroz, Mateusz Sikora, Jürgen Köfinger, Benjamin Schuler, Markus Zweckstetter, Gerhard Hummer.
      The paradigmatic disordered protein tau plays an important role in neuronal function and neurodegenerative diseases. To disentangle the factors controlling the balance between functional and disease-associated conformational states, we build a structural ensemble of the tau K18 fragment containing the four pseudorepeat domains involved in both microtubule binding and amyloid fibril formation. We assemble 129-residue-long tau K18 chains with atomic detail from an extensive fragment library constructed with molecular dynamics simulations. We introduce a reweighted hierarchical chain growth (RHCG) algorithm that integrates experimental data reporting on the local structure into the assembly process in a systematic manner. By combining Bayesian ensemble refinement with importance sampling, we obtain well-defined ensembles and overcome the problem of exponentially varying weights in the integrative modeling of long-chain polymeric molecules. The resulting tau K18 ensembles capture nuclear magnetic resonance (NMR) chemical shift and J-coupling measurements. Without further fitting, we achieve very good agreement with measurements of NMR residual dipolar couplings. The good agreement with experimental measures of global structure such as single-molecule Förster resonance energy transfer (FRET) efficiencies is improved further by ensemble refinement. By comparing wild-type and mutant ensembles, we show that pathogenic single-point P301L, P301S, and P301T mutations shift the population from the turn-like conformations of the functional microtubule-bound state to the extended conformations of disease-associated tau fibrils. RHCG thus provides us with an atomically detailed view of the population equilibrium between functional and aggregation-prone states of tau K18, and demonstrates that global structural characteristics of this intrinsically disordered protein emerge from its local structure.
    DOI:  https://doi.org/10.1021/jacsau.1c00536
  3. J Chem Theory Comput. 2022 Apr 04.
    Improving Martini 3 for Disordered and Multidomain Proteins.
    F Emil Thomasen, Francesco Pesce, Mette Ahrensback Roesgaard, Giulio Tesei, Kresten Lindorff-Larsen.
      Coarse-grained molecular dynamics simulations are a useful tool to determine conformational ensembles of proteins. Here, we show that the coarse-grained force field Martini 3 underestimates the global dimensions of intrinsically disordered proteins (IDPs) and multidomain proteins when compared with small-angle X-ray scattering (SAXS) data and that increasing the strength of protein-water interactions favors more expanded conformations. We find that increasing the strength of interactions between protein and water by ca. 10% results in improved agreement with the SAXS data for IDPs and multidomain proteins. We also show that this correction results in a more accurate description of self-association of IDPs and folded proteins and better agreement with paramagnetic relaxation enhancement data for most IDPs. While simulations with this revised force field still show deviations to experiments for some systems, our results suggest that it is overall a substantial improvement for coarse-grained simulations of soluble proteins.
    DOI:  https://doi.org/10.1021/acs.jctc.1c01042
  4. iScience. 2022 Apr 15. 25(4): 104099
    Long-range structural preformation in yes-associated protein precedes encounter complex formation with TEAD.
    Michael Feichtinger, Andreas Beier, Mario Migotti, Matthias Schmid, Fedir Bokhovchuk, Patrick Chène, Robert Konrat.
      Yes-associated protein (YAP) is a partly intrinsically disordered protein (IDP) that plays a major role as the downstream element of the Hippo pathway. Although the structures of the complex between TEA domain transcription factors (TEADs) and the TEAD-binding domain of YAP are already well characterized, its apo state and the binding mechanism with TEADs are still not clearly defined. Here we characterize via a combination of different NMR approaches with site-directed mutagenesis and affinity measurements the intrinsically disordered solution state of apo YAP. Our results provide evidence that the apo state of YAP adopts several compact conformations that may facilitate the formation of the YAP:TEAD complex. The interplay between local secondary structure element preformation and long-range co-stabilization of these structured elements precedes the encounter complex formation with TEAD and we, therefore, propose that TEAD binding proceeds largely via conformational selection of the preformed compact substates displaying at least nanosecond lifetimes.
    Keywords:  Biochemistry; Cell biology; Structural biology
    DOI:  https://doi.org/10.1016/j.isci.2022.104099
  5. J Chem Inf Model. 2022 Apr 06.
    Dynamic Heterogeneity at the Interface of an Intrinsically Disordered Peptide.
    Souvik Mondal, Krishna Prasad Ghanta, Sanjoy Bandyopadhyay.
      It is believed that water around an intrinsically disordered protein or peptide (IDP) in an aqueous environment plays an important role in guiding its conformational properties and aggregation behavior. However, despite its importance, only a handful of studies exploring the correlation between the conformational motions of an IDP and the microscopic properties of water at its surface are reported. Attempts have been made in this work to study the dynamic properties of water present in the vicinity of α-synuclein, an IDP associated with Parkinson's disease (PD). Room temperature molecular dynamics (MD) simulations of eight α-synuclein1-95 peptides with a wide range of initial conformations have been carried out in aqueous media. The calculations revealed that due to solid-like caging motions, the translational and rotational mobility of water molecules near the surfaces of the peptide repeat unit segments R1 to R7 are significantly restricted. A small degree of dynamic heterogeneity in the hydration environment around the repeat units has been observed with water near the hydrophobic R6 unit exhibiting relatively more restricted diffusivity. The time scales involving the overall structural relaxations of peptide-water and water-water hydrogen bonds near the peptide have been found to be correlated with the time scale of diffusion of the interfacial water molecules. We believe that the relatively more hindered dynamic environment near R6 can help create water-mediated contacts centered around R6 between peptide monomers at a higher concentration, thereby enhancing the early stages of peptide aggregation.
    DOI:  https://doi.org/10.1021/acs.jcim.2c00019
  6. J Biol Chem. 2022 Apr 04. pii: S0021-9258(22)00342-8. [Epub ahead of print] 101902
    The small aromatic compound SynuClean-D inhibits the aggregation and seeded polymerization of multiple α-synuclein strains.
    Samuel Peña-Díaz, Jordi Pujols, Eftychia Vasili, Francisca Pinheiro, Jaime Santos, Zoe Manglano-Artuñedo, Tiago F Outeiro, Salvador Ventura.
      Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra, as well as the accumulation of intra-neuronal proteinaceous inclusions known as Lewy bodies and Lewy neurites. The major protein component of Lewy inclusions is the intrinsically disordered protein α-Synuclein (α-Syn), which can adopt diverse amyloid structures. Different conformational strains of α-Syn have been proposed to be related to the onset of distinct synucleinopathies; however, how specific amyloid fibrils cause distinctive pathological traits is not clear. Here, we generated three different α-Syn amyloid conformations at different pH and salt concentrations and analyzed the activity of SynuClean-D (SC-D), a small aromatic molecule, on these strains. We show that incubation of α-Syn with SC-D reduced the formation of aggregates and the seeded polymerization of α-Syn in all cases. Moreover, we found that SC-D exhibited a general fibril disaggregation activity. Finally, we demonstrate that treatment with SC-D also reduced strain-specific intracellular accumulation of phosphorylated α-Syn inclusions. Taken together, we conclude that SC-D may be a promising hit compound to inhibit polymorphic α-Syn aggregation.
    Keywords:  Parkinson’s disease; aggregation inhibitor; amyloid; small molecule; strains; synucleinopathies; α-synuclein
    DOI:  https://doi.org/10.1016/j.jbc.2022.101902
  7. Int J Biol Macromol. 2022 Apr 02. pii: S0141-8130(22)00695-X. [Epub ahead of print]
    In vitro characterization and molecular dynamics simulation reveal mechanism of 14-3-3ζ regulated phase separation of the tau protein.
    Yue Han, Haiqiong Ye, Ping Li, Yifan Zeng, Jing Yang, Meng Gao, Zhengding Su, Yongqi Huang.
      As a major microtubule-associated protein, tau is involved in the assembly of microtubules in the central nervous system. However, under pathological conditions tau assembles into amyloid filaments. Liquid droplets formed by liquid-liquid phase separation (LLPS) are a recently identified assembly state of tau and may have a major effect on the physiological function of tau and the formation of tau aggregates. 14-3-3 proteins are ubiquitously expressed in various tissues and regulate a wide variety of biological processes. In this work, we demonstrate that 14-3-3ζ is recruited into tau droplets and regulates tau LLPS by in vitro assays. While the mobility of tau molecules inside the droplets is not affected in the presence of 14-3-3ζ, the amount and size of droplets can vary significantly. Mechanistic studies reveal that 14-3-3ζ regulates tau LLPS by electrostatic interactions and hydrophobic interactions with the proline-rich domain and the microtubule-binding domain of tau. Surprisingly, the disordered C-terminal tail rather than the amphipathic binding groove of 14-3-3ζ plays a key role. Our findings not only provide a novel dimension to understand the interactions between 14 and 3-3 proteins and tau, but also suggest that 14-3-3 proteins may play an important role in regulating the LLPS of their binding partners.
    Keywords:  Liquid droplet; Neurodegenerative diseases; Protein aggregation; Protein−protein interaction
    DOI:  https://doi.org/10.1016/j.ijbiomac.2022.03.215
  8. PLoS Comput Biol. 2022 Apr 05. 18(4): e1009578
    Implementation of residue-level coarse-grained models in GENESIS for large-scale molecular dynamics simulations.
    Cheng Tan, Jaewoon Jung, Chigusa Kobayashi, Diego Ugarte La Torre, Shoji Takada, Yuji Sugita.
      Residue-level coarse-grained (CG) models have become one of the most popular tools in biomolecular simulations in the trade-off between modeling accuracy and computational efficiency. To investigate large-scale biological phenomena in molecular dynamics (MD) simulations with CG models, unified treatments of proteins and nucleic acids, as well as efficient parallel computations, are indispensable. In the GENESIS MD software, we implement several residue-level CG models, covering structure-based and context-based potentials for both well-folded biomolecules and intrinsically disordered regions. An amino acid residue in protein is represented as a single CG particle centered at the Cα atom position, while a nucleotide in RNA or DNA is modeled with three beads. Then, a single CG particle represents around ten heavy atoms in both proteins and nucleic acids. The input data in CG MD simulations are treated as GROMACS-style input files generated from a newly developed toolbox, GENESIS-CG-tool. To optimize the performance in CG MD simulations, we utilize multiple neighbor lists, each of which is attached to a different nonbonded interaction potential in the cell-linked list method. We found that random number generations for Gaussian distributions in the Langevin thermostat are one of the bottlenecks in CG MD simulations. Therefore, we parallelize the computations with message-passing-interface (MPI) to improve the performance on PC clusters or supercomputers. We simulate Herpes simplex virus (HSV) type 2 B-capsid and chromatin models containing more than 1,000 nucleosomes in GENESIS as examples of large-scale biomolecular simulations with residue-level CG models. This framework extends accessible spatial and temporal scales by multi-scale simulations to study biologically relevant phenomena, such as genome-scale chromatin folding or phase-separated membrane-less condensations.
    DOI:  https://doi.org/10.1371/journal.pcbi.1009578
  9. Front Cell Dev Biol. 2022 ;10 840256
    Emerging Roles for Phase Separation of RNA-Binding Proteins in Cellular Pathology of ALS.
    Katarina Milicevic, Branislava Rankovic, Pavle R Andjus, Danijela Bataveljic, Dragomir Milovanovic.
      Liquid-liquid phase separation (LLPS) is emerging as a major principle for the mesoscale organization of proteins, RNAs, and membrane-bound organelles into biomolecular condensates. These condensates allow for rapid cellular responses to changes in metabolic activities and signaling. Nowhere is this regulation more important than in neurons and glia, where cellular physiology occurs simultaneously on a range of time- and length-scales. In a number of neurodegenerative diseases, such as Amyotrophic Lateral Sclerosis (ALS), misregulation of biomolecular condensates leads to the formation of insoluble aggregates-a pathological hallmark of both sporadic and familial ALS. Here, we summarize how the emerging knowledge about the LLPS of ALS-related proteins corroborates with their aggregation. Understanding the mechanisms that lead to protein aggregation in ALS and how cells respond to these aggregates promises to open new directions for drug development.
    Keywords:  FUS; RNP aggregates; TDP-43; amyotrophic lateral sclerosis; liquid-liquid phase separation; neurons; stress granule
    DOI:  https://doi.org/10.3389/fcell.2022.840256
  10. Methods. 2022 Mar 31. pii: S1046-2023(22)00085-8. [Epub ahead of print]
    Resources for computational prediction of intrinsic disorder in proteins.
    Lukasz Kurgan.
      With over 40 years of research, researchers in the intrinsic disorder prediction field developed over 100 computational predictors. This review offers a holistic perspective of this field by highlighting accurate and popular disorder predictors and introducing a wide range of practical resources that support collection, interpretation and application of disorder predictions. These resources include meta webservers that expedite collection of multiple disorder predictions, large databases of pre-computed disorder predictions that ease collection of predictions particularly for large datasets of proteins, and modern quality assessment tools. The latter methods facilitate identification of accurate predictions in a specific protein sequence, reducing uncertainty associated to the use of the putative disorder. Altogether, we review eleven predictors, four meta webservers, three databases and two quality assessment tools, all of which are conveniently available online. We also offer a perspective on future developments of the disorder prediction and the quality assessment tools. The availability of this comprehensive toolbox of useful resources should stimulate further growth in the application of the disorder predictions across many areas including rational drug design, systems medicine, structural bioinformatics and structural genomics.
    Keywords:  Intrinsic disorder; Intrinsically disordered proteins; Machine learning; Prediction; Protein function; Webserver
    DOI:  https://doi.org/10.1016/j.ymeth.2022.03.018
  11. EMBO Rep. 2022 Apr 04. e54105
    PLETHORA-WOX5 interaction and subnuclear localization control Arabidopsis root stem cell maintenance.
    Rebecca C Burkart, Vivien I Strotmann, Gwendolyn K Kirschner, Abdullah Akinci, Laura Czempik, Anika Dolata, Alexis Maizel, Stefanie Weidtkamp-Peters, Yvonne Stahl.
      Maintenance and homeostasis of the stem cell niche (SCN) in the Arabidopsis root is essential for growth and development of all root cell types. The SCN is organized around a quiescent center (QC) maintaining the stemness of cells in direct contact. The key transcription factors (TFs) WUSCHEL-RELATED HOMEOBOX 5 (WOX5) and PLETHORAs (PLTs) are expressed in the SCN where they maintain the QC and regulate distal columella stem cell (CSC) fate. Here, we describe the concerted mutual regulation of the key TFs WOX5 and PLTs on a transcriptional and protein interaction level. Additionally, by applying a novel SCN staining method, we demonstrate that both WOX5 and PLTs regulate root SCN homeostasis as they control QC quiescence and CSC fate interdependently. Moreover, we uncover that some PLTs, especially PLT3, contain intrinsically disordered prion-like domains (PrDs) that are necessary for complex formation with WOX5 and its recruitment to subnuclear microdomains/nuclear bodies (NBs) in the CSCs. We propose that this partitioning of PLT-WOX5 complexes to NBs, possibly by phase separation, is important for CSC fate determination.
    Keywords:  differentiation; nuclear bodies; prion-like domains; root stem cells; transcription factor complexes
    DOI:  https://doi.org/10.15252/embr.202154105
  12. J Biol Chem. 2022 Apr 01. pii: S0021-9258(22)00338-6. [Epub ahead of print] 101898
    Functional inhibition of c-Myc using novel inhibitors identified through "hot spot" targeting.
    Ashutosh Singh, Prateek Kumar, Sailu Sarvagalla, Taniya Bharadwaj, Namyashree Nayak, Mohane Selvaraj Coumar, Rajanish Giri, Neha Garg.
      Protein-protein interactions drive various biological processes in healthy as well as disease states. The transcription factor c-Myc plays a crucial role in maintaining cellular homeostasis and its deregulated expression is linked to various human cancers; therefore, it can be considered a viable target for cancer therapeutics. However, the structural heterogeneity of c-Myc due to its disordered nature poses a major challenge to drug discovery. In the present study, we used an in-silico alanine scanning mutagenesis approach to identify "hot-spot" residues within the c-Myc/Myc-associated factor X (MAX) interface, which is highly disordered and has not yet been systematically analyzed for potential small molecule binding sites. We then used the information gained from this analysis to screen potential inhibitors using a conformation ensemble approach. The fluorescence-based biophysical experiments showed that the identified hit molecules displayed non-covalent interactions with these hot-spot residues, and further cell-based experiments showed substantial in vitro potency against diverse c-Myc-expressing cancer/stem cells by deregulating c-Myc activity. These biophysical and computational studies demonstrated stable binding of the hit compounds with the disordered c-Myc protein. Collectively, our data indicated effective drug targeting of the disordered c-Myc protein via the determination of hot-spot residues in the c-Myc/MAX heterodimer.
    Keywords:  c-Myc; cancer stem cells; compound inhibitors; confirmation ensemble; drug discovery; hot spot
    DOI:  https://doi.org/10.1016/j.jbc.2022.101898
  13. Nucleic Acids Res. 2022 Apr 07. pii: gkac233. [Epub ahead of print]
    A histidine cluster determines YY1-compartmentalized coactivators and chromatin elements in phase-separated enhancer clusters.
    Wenmeng Wang, Shiyao Qiao, Guangyue Li, Jiahui Cheng, Cuicui Yang, Chen Zhong, Daniel B Stovall, Jinming Shi, Chunbo Teng, Dangdang Li, Guangchao Sui.
      As an oncogenic transcription factor, Yin Yang 1 (YY1) regulates enhancer and promoter connection. However, gaps still exist in understanding how YY1 coordinates coactivators and chromatin enhancer elements to assemble enhancers and super-enhancers. Here, we demonstrate that a histidine cluster in YY1's transactivation domain is essential for its formation of phase separation condensates, which can be extended to additional proteins. The histidine cluster is also required for YY1-promoted cell proliferation, migration, clonogenicity and tumor growth. YY1-rich nuclear puncta contain coactivators EP300, BRD4, MED1 and active RNA polymerase II, and colocalize with histone markers of gene activation, but not that of repression. Furthermore, YY1 binds to the consensus motifs in the FOXM1 promoter to activate its expression. Wild-type YY1, but not its phase separation defective mutant, connects multiple enhancer elements and the FOXM1 promoter to form an enhancer cluster. Consistently, fluorescent in situ hybridization (FISH) assays reveal the colocalization of YY1 puncta with both the FOXM1 gene locus and its nascent RNA transcript. Overall, this study demonstrates that YY1 activates target gene expression through forming liquid-liquid phase separation condensates to compartmentalize both coactivators and enhancer elements, and the histidine cluster of YY1 plays a determinant role in this regulatory mechanism.
    DOI:  https://doi.org/10.1093/nar/gkac233
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