bims-indpro Biomed News
on Intrinsically disordered proteins
Issue of 2022–07–10
fourteen papers selected by
Sara Mingu, Johannes Gutenberg University
  1. Intrinsically disordered proteins and proteins with intrinsically disordered regions in neurodegenerative diseases.
  2. Exploring Manually Curated Annotations of Intrinsically Disordered Proteins with DisProt.
  3. Plasticity of Membrane Binding by the Central Region of α-Synuclein.
  4. Negative Thermal Expansion and Disorder-to-Order Collapse of an Intrinsically Disordered Protein under Marginally Denaturing Conditions.
  5. Intrinsically disordered CO2 sensors.
  6. Editorial: Fuzzy Interactions: Many Facets of Protein Binding.
  7. The Multivalent Polyampholyte Domain of Nst1, a P-Body-Associated Saccharomyces cerevisiae Protein, Provides a Platform for Interacting with P-Body Components.
  8. Genetically Encoded Aryl Alkyne for Raman Spectral Imaging of Intracellular α-Synuclein Fibrils.
  9. Role of Helical Structure in MBP Immunodominant Peptides for Efficient IgM Antibody Recognition in Multiple Sclerosis.
  10. The C-Terminal Domain of Y-Box Binding Protein 1 Exhibits Structure-Specific Binding to Poly(ADP-Ribose), Which Regulates PARP1 Activity.
  11. An entropic safety catch controls Hepatitis C virus entry and antibody resistance.
  12. Phase-separating RNA-binding proteins form heterogeneous distributions of clusters in subsaturated solutions.
  13. Liquid-liquid phase separation of α-Synuclein: A new mechanistic insight for α-Synuclein aggregation associated with Parkinson's disease pathogenesis.
  14. Variation of Structural and Dynamical Flexibility of Myelin Basic Protein in Response to Guanidinium Chloride.
  1. Biophys Rev. 2022 Jun;14(3): 679-707
    Intrinsically disordered proteins and proteins with intrinsically disordered regions in neurodegenerative diseases.
    Orkid Coskuner-Weber, Ozan Mirzanli, Vladimir N Uversky.
      Many different intrinsically disordered proteins and proteins with intrinsically disordered regions are associated with neurodegenerative diseases. These types of proteins including amyloid-β, tau, α-synuclein, CHCHD2, CHCHD10, and G-protein coupled receptors are increasingly becoming evaluated as potential drug targets in the pharmaceutical-based treatment approaches. Here, we focus on the neurobiology of this class of proteins, which lie at the center of numerous neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases, Huntington's disease, amyotrophic lateral sclerosis, frontotemporal dementia, Charcot-Marie-Tooth diseases, spinal muscular atrophy, and mitochondrial myopathy. Furthermore, we discuss the current treatment design strategies involving intrinsically disordered proteins and proteins with intrinsically disordered regions in neurodegenerative diseases. In addition, we emphasize that although the G-protein coupled receptors are traditionally investigated using structural biology-based models and approaches, current studies show that these receptors are proteins with intrinsically disordered regions and therefore they require new ways for their analysis.
    Keywords:  Drug design; Intrinsically disordered proteins; Neurodegeneration; Pathology; Proteins with intrinsically disordered regions
    DOI:  https://doi.org/10.1007/s12551-022-00968-0
  2. Curr Protoc. 2022 Jul;2(7): e484
    Exploring Manually Curated Annotations of Intrinsically Disordered Proteins with DisProt.
    Federica Quaglia, András Hatos, Edoardo Salladini, Damiano Piovesan, Silvio C E Tosatto.
      DisProt is the major repository of manually curated data for intrinsically disordered proteins collected from the literature. Although lacking a stable three-dimensional structure under physiological conditions, intrinsically disordered proteins carry out a plethora of biological functions, some of them directly arising from their flexible nature. A growing number of scientific studies have been published during the last few decades to shed light on their unstructured state, their binding modes, and their functions. DisProt makes use of a team of expert biocurators to provide up-to-date annotations of intrinsically disordered proteins from the literature, making them available to the scientific community. Here we present a comprehensive description on how to use DisProt in different contexts and provide a detailed explanation of how to explore and interpret manually curated annotations of intrinsically disordered proteins. We describe how to search DisProt annotations, both using the web interface and the API for programmatic access. Finally, we explain how to visualize and interpret a DisProt entry, the SARS-CoV-2 Nucleoprotein, characterized by the presence of unstructured N-terminal and C-terminal regions and a flexible linker. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Performing a search in DisProt Support Protocol 1: Downloading options Support Protocol 2: Programmatic access with DisProt REST API Basic Protocol 2: Exploring the DisProt Ontology page Basic Protocol 3: Visualizing and interpreting DisProt entries-the SARS-CoV-2 Nucleoprotein use case.
    Keywords:  DisProt; IDPs; biocuration; database; intrinsically disordered proteins
    DOI:  https://doi.org/10.1002/cpz1.484
  3. Front Mol Biosci. 2022 ;9 857217
    Plasticity of Membrane Binding by the Central Region of α-Synuclein.
    Carlos Navarro-Paya, Maximo Sanz-Hernandez, Alfonso De Simone.
      Membrane binding by α-synuclein (αS), an intrinsically disordered protein whose aggregation is associated with Parkinson's disease, is a key step in determining its biological properties under both physiological and pathological conditions. Upon membrane interaction, αS retains a partial level of structural disorder despite acquiring α-helical content. In the membrane-bound state, the equilibrium between the helical-bound and disordered-detached states of the central region of αS (residues 65-97) has been involved in a double-anchor mechanism that promotes the clustering of synaptic vesicles. Herein, we investigated the underlying molecular bases of this equilibrium using enhanced coarse-grained molecular dynamics simulations. The results enabled clarifying the conformational dependencies of the membrane affinity by this protein region that, in addition to playing a role in physiological membrane binding, has key relevance for the aggregation of αS and the mechanisms of the toxicity of the resulting assemblies.
    Keywords:  coarse-grained simulations; double-anchor mechanism; intrinsically disordered proteins; membrane binding; vesicle clustering; α-synuclein
    DOI:  https://doi.org/10.3389/fmolb.2022.857217
  4. J Phys Chem B. 2022 Jul 05.
    Negative Thermal Expansion and Disorder-to-Order Collapse of an Intrinsically Disordered Protein under Marginally Denaturing Conditions.
    Abani K Bhuyan.
      Recent work with intrinsically disordered proteins (IDPs) has projected a myriad of their survival instincts based mainly on the total charge content, the abundance of polar residues, and the paucity of hydrophobic amino acids. This work uses a plant IDP AtPP16-1 (Arabidopsis thaliana phloem protein class 16-1), whose solution NMR structure was determined by us recently, to show legitimate negative thermal expansion (NTE) of the native state. The thermal expansion continues to be negative even as the tertiary structure is perturbed by ultralow levels of urea up to 0.4 M. The NTE of these subdenatured states is called apparent NTE because they are prone to undergo conformational changes with temperature. Hydrodynamic shrinkage of the NTE IDP is also observed by dynamic light scattering (DLS) and NMR-measured global rotational correlation time (τc). The protein with denatured tertiary structure but otherwise preserved native-state secondary structure collapses to a dynamically rigid state. The data are mainly based on thermal coefficients of chemical shift and nuclear relaxation measured by heteronuclear NMR. The hydrodynamic shrinkage and collapse under marginally varying solvent compositions that may arise from unstable tertiary structure and dynamic disorder of chain segments across the backbone could be a generic property of IDPs.
    DOI:  https://doi.org/10.1021/acs.jpcb.2c03386
  5. Nat Cell Biol. 2022 Jul 05.
    Intrinsically disordered CO2 sensors.
    Ryan J Emenecker, Alex S Holehouse.
      
    DOI:  https://doi.org/10.1038/s41556-022-00957-1
  6. Front Mol Biosci. 2022 ;9 947215
    Editorial: Fuzzy Interactions: Many Facets of Protein Binding.
    Damiano Piovesan, Miguel Arbesú, Monika Fuxreiter, Miquel Pons.
      
    Keywords:  amyloids; chaperones; conformational ensembles; disordered drug targets; fuzzy complexes; intrinsically disordered proteins; thermodynamics; transcription regulation
    DOI:  https://doi.org/10.3389/fmolb.2022.947215
  7. Int J Mol Sci. 2022 Jul 02. pii: 7380. [Epub ahead of print]23(13):
    The Multivalent Polyampholyte Domain of Nst1, a P-Body-Associated Saccharomyces cerevisiae Protein, Provides a Platform for Interacting with P-Body Components.
    Yoon-Jeong Choi, Yujin Lee, Yuxi Lin, Yunseok Heo, Young-Ho Lee, Kiwon Song.
      The condensation of nuclear promyelocytic leukemia bodies, cytoplasmic P-granules, P-bodies (PBs), and stress granules is reversible and dynamic via liquid-liquid phase separation. Although each condensate comprises hundreds of proteins with promiscuous interactions, a few key scaffold proteins are required. Essential scaffold domain sequence elements, such as poly-Q, low-complexity regions, oligomerizing domains, and RNA-binding domains, have been evaluated to understand their roles in biomolecular condensation processes. However, the underlying mechanisms remain unclear. We analyzed Nst1, a PB-associated protein that can intrinsically induce PB component condensations when overexpressed. Various Nst1 domain deletion mutants with unique sequence distributions, including intrinsically disordered regions (IDRs) and aggregation-prone regions, were constructed based on structural predictions. The overexpression of Nst1 deletion mutants lacking the aggregation-prone domain (APD) significantly inhibited self-condensation, implicating APD as an oligomerizing domain promoting self-condensation. Remarkably, cells overexpressing the Nst1 deletion mutant of the polyampholyte domain (PD) in the IDR region (Nst1∆PD) rarely accumulate endogenous enhanced green fluorescent protein (EGFP)-tagged Dcp2. However, Nst1∆PD formed self-condensates, suggesting that Nst1 requires PD to interact with Dcp2, regardless of its self-condensation. In Nst1∆PD-overexpressing cells treated with cycloheximide (CHX), Dcp2, Xrn1, Dhh1, and Edc3 had significantly diminished condensation compared to those in CHX-treated Nst1-overexpressing cells. These observations suggest that the PD of the IDR in Nst1 functions as a hub domain interacting with other PB components.
    Keywords:  Nst1; P-body; Saccharomyces cerevisiae; aggregation-prone domain; liquid–liquid phase separation; polyampholyte domain
    DOI:  https://doi.org/10.3390/ijms23137380
  8. J Mol Biol. 2022 Jul 02. pii: S0022-2836(22)00308-4. [Epub ahead of print] 167716
    Genetically Encoded Aryl Alkyne for Raman Spectral Imaging of Intracellular α-Synuclein Fibrils.
    Matthew D Watson, Jennifer C Lee.
      α-Synuclein (α-syn) is an intrinsically disordered protein involved in a group of diseases collectively termed synucleinopathies, characterized by the aggregation of α-syn to form insoluble, β-sheet-rich amyloid fibrils. Amyloid fibrils are thought to contribute to disease progression through cell-to-cell transmission, templating and propagating intracellular amyloid formation. Raman spectral imaging offers a direct characterization of protein secondary structure via the amide-I backbone vibration; however, specific detection of α-syn conformational changes against the background of other cellular components presents a challenge. Here, we demonstrate the ability to unambiguously identify cellularly internalized α-syn fibrils by coupling Raman spectral imaging with the use of a genetically encoded aryl alkyne, 4-ethynyl-l-phenylalanine (FCC), through amber codon suppression. The alkyne stretch (C≡C) of FCC provides a spectrally unique molecular vibration without interference from native biomolecules. Cellular uptake of FCC-α-syn fibrils formed in vitro was visualized in cultured human SH-SY5Y neuroblastoma cells by Raman spectral imaging. Fibrils appear as discrete cytosolic clusters of varying sizes, found often at the cellular periphery. Raman spectra of internalized fibrils exhibit frequency shifts and spectral narrowing relative to in vitro fibrils, highlighting the environmental sensitivity of the alkyne vibration. Interestingly, spectral analysis reveals variations in lipid and protein recruitment to these aggregates, and in some cases, secondary structural changes in the fibrils are observed. This work sets the groundwork for future Raman spectroscopic investigations using a similar approach of an evolved aminoacyl-tRNA synthetase/tRNA pair to incorporate FCC into endogenous amyloidogenic proteins to monitor their aggregation in cells.
    Keywords:  4-ethynylphenylalanine; Parkinson’s disease; SH-SY5Y; amyloid; unnatural amino acid
    DOI:  https://doi.org/10.1016/j.jmb.2022.167716
  9. Front Chem. 2022 ;10 885180
    Role of Helical Structure in MBP Immunodominant Peptides for Efficient IgM Antibody Recognition in Multiple Sclerosis.
    Agnieszka Staśkiewicz, Michael Quagliata, Feliciana Real-Fernandez, Francesca Nuti, Roberta Lanzillo, Vincenzo Brescia-Morra, Hendrik Rusche, Michal Jewginski, Alfonso Carotenuto, Diego Brancaccio, Rina Aharoni, Ruth Arnon, Paolo Rovero, Rafal Latajka, Anna Maria Papini.
      The involvement of Myelin Basic Protein (MBP) in Multiple Sclerosis (MS) has been widely discussed in the literature. This intrinsically disordered protein has an interesting α-helix motif, which can be considered as a conformational epitope. In this work we investigate the importance of the helical structure in antibody recognition by MBP peptides of different lengths. Firstly, we synthesized the peptide MBP (81-106) (1) and observed that its elongation at both N- and C-termini, to obtain the peptide MBP (76-116) (2) improves IgM antibody recognition in SP-ELISA, but destabilizes the helical structure. Conversely, in competitive ELISA, MBP (81-106) (1) is recognized more efficiently by IgM antibodies than MBP (76-116) (2), possibly thanks to its more stable helical structure observed in CD and NMR conformational experiments. These results are discussed in terms of different performances of peptide antigens in the two ELISA formats tested.
    Keywords:  NMR; circular dichroism; immune response; multiple sclerosis; myelin basic protein antigen; peptide-antigen based ELISA; synthetic helical peptides
    DOI:  https://doi.org/10.3389/fchem.2022.885180
  10. Front Cell Dev Biol. 2022 ;10 831741
    The C-Terminal Domain of Y-Box Binding Protein 1 Exhibits Structure-Specific Binding to Poly(ADP-Ribose), Which Regulates PARP1 Activity.
    Konstantin N Naumenko, Mariya V Sukhanova, Loic Hamon, Tatyana A Kurgina, Rashid O Anarbaev, Aswin Mangerich, David Pastré, Olga I Lavrik.
      Y-box-binding protein 1 (YB-1) is a multifunctional protein involved in the regulation of gene expression. Recent studies showed that in addition to its role in the RNA and DNA metabolism, YB-1 is involved in the regulation of PARP1 activity, which catalyzes poly(ADP-ribose) [PAR] synthesis under genotoxic stress through auto-poly(ADP-ribosyl)ation or protein trans-poly(ADP-ribosyl)ation. Nonetheless, the exact mechanism by which YB-1 regulates PAR synthesis remains to be determined. YB-1 contains a disordered Ala/Pro-rich N-terminal domain, a cold shock domain, and an intrinsically disordered C-terminal domain (CTD) carrying four clusters of positively charged amino acid residues. Here, we examined the functional role of the disordered CTD of YB-1 in PAR binding and in the regulation of PARP1-driven PAR synthesis in vitro. We demonstrated that the rate of PARP1-dependent synthesis of PAR is higher in the presence of YB-1 and is tightly controlled by the interaction between YB-1 CTD and PAR. Moreover, YB-1 acts as an effective cofactor in the PAR synthesis catalyzed by the PARP1 point mutants that generate various PAR polymeric structures, namely, short hypo- or hyperbranched polymers. We showed that either a decrease in chain length or an increase in branching frequency of PAR affect its binding affinity for YB-1 and YB-1-mediated stimulation of PARP1 enzymatic activity. These results provide important insight into the mechanism underlying the regulation of PARP1 activity by PAR-binding proteins containing disordered regions with clusters of positively charged amino acid residues, suggesting that YB-1 CTD-like domains may be considered PAR "readers" just as other known PAR-binding modules.
    Keywords:  PARP1; Y-box binding protein 1; disordered C-terminal domain; poly(ADP-ribose); trans-poly(ADP-ribosyl) ation
    DOI:  https://doi.org/10.3389/fcell.2022.831741
  11. Elife. 2022 Jul 07. pii: e71854. [Epub ahead of print]11
    An entropic safety catch controls Hepatitis C virus entry and antibody resistance.
    Lenka Stejskal, Mphatso D Kalemera, Charlotte B Lewis, Machaela Palor, Lucas Walker, Tina Daviter, William D Lees, David S Moss, Myrto Kremyda-Vlachou, Zisis Zisis Kozlakidis, Giulia Gallo, Dalan Bailey, William Rosenberg, Christopher J R Illingworth, Adrian J Shepherd, Joe Grove.
      E1 and E2 (E1E2), the fusion proteins of Hepatitis C Virus (HCV), are unlike that of any other virus yet described, and the detailed molecular mechanisms of HCV entry/fusion remain unknown. Hypervariable region-1 (HVR-1) of E2 is a putative intrinsically disordered protein tail. Here, we demonstrate that HVR-1 has an autoinhibitory function that suppresses the activity of E1E2 on free virions; this is dependent on its conformational entropy. Thus, HVR-1 is akin to a safety catch that prevents premature triggering of E1E2 activity. Crucially, this mechanism is turned off by host receptor interactions at the cell surface to allow entry. Mutations that reduce conformational entropy in HVR-1, or genetic deletion of HVR-1, turn off the safety catch to generate hyper-reactive HCV that exhibits enhanced virus entry but is thermally unstable and acutely sensitive to neutralising antibodies. Therefore, the HVR-1 safety catch controls the efficiency of virus entry and maintains resistance to neutralising antibodies. This discovery provides an explanation for the ability of HCV to persist in the face of continual immune assault and represents a novel regulatory mechanism that is likely to be found in other viral fusion machinery.
    Keywords:  infectious disease; microbiology; molecular biophysics; structural biology; viruses
    DOI:  https://doi.org/10.7554/eLife.71854
  12. Proc Natl Acad Sci U S A. 2022 Jul 12. 119(28): e2202222119
    Phase-separating RNA-binding proteins form heterogeneous distributions of clusters in subsaturated solutions.
    Mrityunjoy Kar, Furqan Dar, Timothy J Welsh, Laura T Vogel, Ralf Kühnemuth, Anupa Majumdar, Georg Krainer, Titus M Franzmann, Simon Alberti, Claus A M Seidel, Tuomas P J Knowles, Anthony A Hyman, Rohit V Pappu.
      Macromolecular phase separation is thought to be one of the processes that drives the formation of membraneless biomolecular condensates in cells. The dynamics of phase separation are thought to follow the tenets of classical nucleation theory, and, therefore, subsaturated solutions should be devoid of clusters with more than a few molecules. We tested this prediction using in vitro biophysical studies to characterize subsaturated solutions of phase-separating RNA-binding proteins with intrinsically disordered prion-like domains and RNA-binding domains. Surprisingly, and in direct contradiction to expectations from classical nucleation theory, we find that subsaturated solutions are characterized by the presence of heterogeneous distributions of clusters. The distributions of cluster sizes, which are dominated by small species, shift continuously toward larger sizes as protein concentrations increase and approach the saturation concentration. As a result, many of the clusters encompass tens to hundreds of molecules, while less than 1% of the solutions are mesoscale species that are several hundred nanometers in diameter. We find that cluster formation in subsaturated solutions and phase separation in supersaturated solutions are strongly coupled via sequence-encoded interactions. We also find that cluster formation and phase separation can be decoupled using solutes as well as specific sets of mutations. Our findings, which are concordant with predictions for associative polymers, implicate an interplay between networks of sequence-specific and solubility-determining interactions that, respectively, govern cluster formation in subsaturated solutions and the saturation concentrations above which phase separation occurs.
    Keywords:  associative polymers; mesoscale clusters; phase separation; sol–gel transitions; stickers and spacers
    DOI:  https://doi.org/10.1073/pnas.2202222119
  13. J Mol Biol. 2022 Jul 01. pii: S0022-2836(22)00305-9. [Epub ahead of print] 167713
    Liquid-liquid phase separation of α-Synuclein: A new mechanistic insight for α-Synuclein aggregation associated with Parkinson's disease pathogenesis.
    Semanti Mukherjee, Arunima Sakunthala, Laxmikant Gadhe, Manisha Poudyal, Ajay Singh Sawner, Pradeep Kadu, Samir K Maji.
      Aberrant aggregation of the misfolded presynaptic protein, α-Synuclein (α-Syn) into Lewy body (LB) and Lewy neuritis (LN) is a major pathological hallmark of Parkinson's disease (PD) and other synucleinopathies. Numerous studies have suggested that prefibrillar and fibrillar species of the misfolded α-Syn aggregates are responsible for cell death in PD pathogenesis. However, the precise molecular events during α-Syn aggregation, especially in the early stages, remain elusive. Emerging evidence has demonstrated that the liquid-liquid phase separation (LLPS) of α-Syn occurs in the nucleation step of α-Syn aggregation, which offers an alternate non-canonical aggregation pathway in the crowded microenvironment. The liquid-like α-Syn droplets gradually undergo an irreversible liquid-to-solid phase transition into amyloid-like hydrogel entrapping oligomers and fibrils. This new mechanism of α-Syn LLPS and gel formation might represent the molecular basis of cellular toxicity associated with PD. This review aims to demonstrate the recent development of α-Syn LLPS, the underlying mechanism along with the microscopic events of aberrant phase transition. This review further discusses how several intrinsic and extrinsic factors regulate the thermodynamics and kinetics of α-Syn LLPS and co-LLPS with other proteins, which might explain the pathophysiology of α-Syn in various neurodegenerative diseases.
    Keywords:  Amyloid; Liquid-liquid phase separation; Liquid-to-solid phase transition; Parkinson’s disease; α-Syn aggregation
    DOI:  https://doi.org/10.1016/j.jmb.2022.167713
  14. Int J Mol Sci. 2022 Jun 23. pii: 6969. [Epub ahead of print]23(13):
    Variation of Structural and Dynamical Flexibility of Myelin Basic Protein in Response to Guanidinium Chloride.
    Luman Haris, Ralf Biehl, Martin Dulle, Aurel Radulescu, Olaf Holderer, Ingo Hoffmann, Andreas M Stadler.
      Myelin basic protein (MBP) is intrinsically disordered in solution and is considered as a conformationally flexible biomacromolecule. Here, we present a study on perturbation of MBP structure and dynamics by the denaturant guanidinium chloride (GndCl) using small-angle scattering and neutron spin-echo spectroscopy (NSE). A concentration of 0.2 M GndCl causes charge screening in MBP resulting in a compact, but still disordered protein conformation, while GndCl concentrations above 1 M lead to structural expansion and swelling of MBP. NSE data of MBP were analyzed using the Zimm model with internal friction (ZIF) and normal mode (NM) analysis. A significant contribution of internal friction was found in compact states of MBP that approaches a non-vanishing internal friction relaxation time of approximately 40 ns at high GndCl concentrations. NM analysis demonstrates that the relaxation rates of internal modes of MBP remain unaffected by GndCl, while structural expansion due to GndCl results in increased amplitudes of internal motions. Within the model of the Brownian oscillator our observations can be rationalized by a loss of friction within the protein due to structural expansion. Our study highlights the intimate coupling of structural and dynamical plasticity of MBP, and its fundamental difference to the behavior of ideal polymers in solution.
    Keywords:  guanidinium chloride; internal protein dynamics; myelin basic protein; neutron spin–echo spectroscopy; normal mode analysis; polymer theory; small-angle scattering
    DOI:  https://doi.org/10.3390/ijms23136969
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