bims-indpro Biomed News
on Intrinsically disordered proteins
Issue of 2022–08–14
thirteen papers selected by
Sara Mingu, Johannes Gutenberg University
  1. Conformational buffering underlies functional selection in intrinsically disordered protein regions.
  2. Predicting Molecular Properties of α-Synuclein Using Force Fields for Intrinsically Disordered Proteins.
  3. ATRANET - Automated generation of transition networks for the structural characterization of intrinsically disordered proteins.
  4. Salt-Induced Transitions in the Conformational Ensembles of Intrinsically Disordered Proteins.
  5. Rational peptide design for regulating liquid-liquid phase separation on the basis of residue-residue contact energy.
  6. Dynamic refolding of OxyS sRNA by the Hfq RNA chaperone.
  7. Evolution of short linear motifs and disordered proteins Topic: yeast as model system to study evolution.
  8. An intrinsically disordered transcription activation domain increases the DNA binding affinity and reduces the specificity of NFκB p50/RelA.
  9. TGA transcription factors-Structural characteristics as basis for functional variability.
  10. Mechanism underlying liquid-to-solid phase transition in fused in sarcoma liquid droplets.
  11. Molecular mechanism for the synchronized electrostatic coacervation and co-aggregation of alpha-synuclein and tau.
  12. NKCC1 and KCC2: Structural insights into phospho-regulation.
  13. Phase Separation Properties in Transcriptional Organization.
  1. Nat Struct Mol Biol. 2022 Aug;29(8): 781-790
    Conformational buffering underlies functional selection in intrinsically disordered protein regions.
    Nicolás S González-Foutel, Juliana Glavina, Wade M Borcherds, Matías Safranchik, Susana Barrera-Vilarmau, Amin Sagar, Alejandro Estaña, Amelie Barozet, Nicolás A Garrone, Gregorio Fernandez-Ballester, Clara Blanes-Mira, Ignacio E Sánchez, Gonzalo de Prat-Gay, Juan Cortés, Pau Bernadó, Rohit V Pappu, Alex S Holehouse, Gary W Daughdrill, Lucía B Chemes.
      Many disordered proteins conserve essential functions in the face of extensive sequence variation, making it challenging to identify the mechanisms responsible for functional selection. Here we identify the molecular mechanism of functional selection for the disordered adenovirus early gene 1A (E1A) protein. E1A competes with host factors to bind the retinoblastoma (Rb) protein, subverting cell cycle regulation. We show that two binding motifs tethered by a hypervariable disordered linker drive picomolar affinity Rb binding and host factor displacement. Compensatory changes in amino acid sequence composition and sequence length lead to conservation of optimal tethering across a large family of E1A linkers. We refer to this compensatory mechanism as conformational buffering. We also detect coevolution of the motifs and linker, which can preserve or eliminate the tethering mechanism. Conformational buffering and motif-linker coevolution explain robust functional encoding within hypervariable disordered linkers and could underlie functional selection of many disordered protein regions.
    DOI:  https://doi.org/10.1038/s41594-022-00811-w
  2. Proteins. 2022 Aug 11.
    Predicting Molecular Properties of α-Synuclein Using Force Fields for Intrinsically Disordered Proteins.
    Kasper B Pedersen, Jose C Flores-Canales, Birgit Schiøtt.
      Independent force field validation is an essential practice to keep track of developments and for performing meaningful Molecular Dynamics simulations. In this work, atomistic force fields for intrinsically disordered proteins (IDP) are tested by simulating the archetypical IDP α-synuclein in solution for 2.5 μs. Four combinations of protein and water force fields were tested: ff19SB/OPC, ff19SB/TIP4P-D, ff03CMAP/TIP4P-D, and a99SB-disp/TIP4P-disp, with four independent repeat simulations for each combination. We compare our simulations to the results of a 73 μs simulation using the a99SB-disp/TIP4P-disp combination, provided by D. E. Shaw Research. From the trajectories, we predict a range of experimental observations of α-synuclein and compare them to literature data. This includes protein radius of gyration and hydration, intramolecular distances, NMR chemical shifts, and 3 J-couplings. Both ff19SB/TIP4P-D and a99SB-disp/TIP4P-disp produce extended conformational ensembles of α-synuclein that agree well with experimental radius of gyration and intramolecular distances while a99SB-disp/TIP4P-disp reproduces a balanced α-synuclein secondary structure content. It was found that ff19SB/OPC and ff03CMAP/TIP4P-D produce overly compact conformational ensembles and show discrepancies in the secondary structure content compared to the experimental data. This article is protected by copyright. All rights reserved.
    Keywords:  Intrinsically Disordered Proteins; Molecular Dynamics Simulations; OPC; TIP4P-D; a99SB-disp; ff03CMAP; ff19SB; α-Synuclein
    DOI:  https://doi.org/10.1002/prot.26409
  3. Methods. 2022 Aug 04. pii: S1046-2023(22)00167-0. [Epub ahead of print]
    ATRANET - Automated generation of transition networks for the structural characterization of intrinsically disordered proteins.
    Moritz Schäffler, Mohammed Khaled, Birgit Strodel.
      Intrinsically disordered proteins (IDPs) do not fold into a unique three-dimensional structure but sample different configurations of different probabilities that further change with the surrounding of the IDPs. The structural heterogeneity and dynamics of IDPs pose a challenge for the characterization of their structures by experimental techniques only. Molecular dynamics (MD) simulations provide a powerful complement to experimental approaches for that purpose. However, MD simulations on the micro- to millisecond timescale generate a lot of data of protein motions, necessitating advanced post-processing techniques to extract the relevant information. Here, we demonstrate how transition networks created from MD trajectories allow revealing the configurational ensemble and structural interconversions of IDPs, using the amyloid-β peptide as example. The construction of transition networks relies on molecular descriptors as input, and we show how the choice of descriptors influences the resulting transition network. The transition networks are generated with the open-source Python script ATRANET, and we explain the usage of ATRANET by providing a detailed workflow and exemplary analysis for amyloid-β, which can be easily generalized to other IDPs and even protein aggregation.
    Keywords:  amyloid-; disorder-to-order transition; intrinsically disordered proteins; molecular dynamics simulations; transition networks
    DOI:  https://doi.org/10.1016/j.ymeth.2022.07.013
  4. J Phys Chem B. 2022 Aug 09.
    Salt-Induced Transitions in the Conformational Ensembles of Intrinsically Disordered Proteins.
    Hiranmay Maity, Lipika Baidya, Govardhan Reddy.
      Salts modulate the behavior of intrinsically disordered proteins (IDPs) and influence the formation of membraneless organelles through liquid-liquid phase separation (LLPS). In low ionic strength solutions, IDP conformations are perturbed by the screening of electrostatic interactions, independent of the salt identity. In this regime, insight into the IDP behavior can be obtained using the theory for salt-induced transitions in charged polymers. However, salt-specific interactions with the charged and uncharged residues, known as the Hofmeister effect, influence IDP behavior in high ionic strength solutions. There is a lack of reliable theoretical models in high salt concentration regimes to predict the salt effect on IDPs. We propose a simulation methodology using a coarse-grained IDP model and experimentally measured water to salt solution transfer free energies of various chemical groups that allowed us to study the salt-specific transitions induced in the IDPs conformational ensemble. We probed the effect of three different monovalent salts on five IDPs belonging to various polymer classes based on charged residue content. We demonstrate that all of the IDPs of different polymer classes behave as self-avoiding walks (SAWs) at physiological salt concentration. In high salt concentrations, the transitions observed in the IDP conformational ensembles are dependent on the salt used and the IDP sequence and composition. Changing the anion with the cation fixed can result in the IDP transition from a SAW-like behavior to a collapsed globule. An important implication of these results is that a suitable salt can be identified to induce condensation of an IDP through LLPS.
    DOI:  https://doi.org/10.1021/acs.jpcb.2c03476
  5. Sci Rep. 2022 Aug 12. 12(1): 13718
    Rational peptide design for regulating liquid-liquid phase separation on the basis of residue-residue contact energy.
    Kiyoto Kamagata, Maulana Ariefai, Hiroto Takahashi, Atsumi Hando, Dwiky Rendra Graha Subekti, Keisuke Ikeda, Atsushi Hirano, Tomoshi Kameda.
      Since liquid-liquid phase separation (LLPS) of proteins is governed by their intrinsically disordered regions (IDRs), it can be controlled by LLPS-regulators that bind to the IDRs. The artificial design of LLPS-regulators based on this mechanism can be leveraged in biological and therapeutic applications. However, the fabrication of artificial LLPS-regulators remains challenging. Peptides are promising candidates for artificial LLPS-regulators because of their ability to potentially bind to IDRs complementarily. In this study, we provide a rational peptide design methodology for targeting IDRs based on residue-residue contact energy obtained using molecular dynamics (MD) simulations. This methodology provides rational peptide sequences that function as LLPS regulators. The peptides designed with the MD-based contact energy showed dissociation constants of 35-280 nM for the N-terminal IDR of the tumor suppressor p53, which are significantly lower than the dissociation constants of peptides designed with the conventional 3D structure-based energy, demonstrating the validity of the present peptide design methodology. Importantly, all of the designed peptides enhanced p53 droplet formation. The droplet-forming peptides were converted to droplet-deforming peptides by fusing maltose-binding protein (a soluble tag) to the designed peptides. Thus, the present peptide design methodology for targeting IDRs is useful for regulating droplet formation.
    DOI:  https://doi.org/10.1038/s41598-022-17829-1
  6. J Mol Biol. 2022 Aug 04. pii: S0022-2836(22)00378-3. [Epub ahead of print] 167776
    Dynamic refolding of OxyS sRNA by the Hfq RNA chaperone.
    Huahuan Cai, Jorjethe Roca, Yu-Fen Zhao, Sarah A Woodson.
      The Sm protein Hfq chaperones small non-coding RNAs (sRNAs) in bacteria, facilitating sRNA regulation of target mRNAs. Hfq acts in part by remodeling the sRNA and mRNA structures, yet the basis for this remodeling activity is not understood. To understand how Hfq remodels RNA, we used single-molecule Förster resonance energy transfer (smFRET) to monitor conformational changes in OxyS sRNA upon Hfq binding. The results show that E. coli Hfq first compacts OxyS, bringing its 5' and 3' ends together. Next, Hfq destabilizes an internal stem-loop in OxyS, allowing the RNA to adopt a more open conformation that is stabilized by a conserved arginine on the rim of Hfq. The frequency of transitions between compact and open conformations depended on interactions with Hfq's flexible C-terminal domain (CTD), being more rapid when the CTD was deleted, and slower when OxyS was bound to Caulobacter crescentus Hfq, which has a shorter and more stable CTD than E. coli Hfq. We propose that the CTDs gate transitions between OxyS conformations that are stabilized by interaction with one or more arginines. These results suggest a general model for how basic residues and intrinsically disordered regions of RNA chaperones act together to refold RNA.
    Keywords:  RNA chaperone; Sm protein; intrinsically disordered protein; single molecule fluorescence microscopy; small RNA
    DOI:  https://doi.org/10.1016/j.jmb.2022.167776
  7. Curr Opin Genet Dev. 2022 Aug 05. pii: S0959-437X(22)00073-9. [Epub ahead of print]76 101964
    Evolution of short linear motifs and disordered proteins Topic: yeast as model system to study evolution.
    Ami G Sangster, Taraneh Zarin, Alan M Moses.
      Evolutionary preservation of protein structure had a major influence on the field of molecular evolution: changes in individual amino acids that did not disrupt protein folding would either have no effect or subtly change the 'lock' so that it could fit a new 'key'. Homology of individual amino acids could be confidently assigned through sequence alignments, and models of evolution could be tested. This view of molecular evolution excluded large regions of proteins that could not be confidently aligned, such as intrinsically disordered regions (IDRs) that do not fold into stable structures. In the last decade, major progress has been made in understanding the evolution of IDRs, much of it facilitated by new experimental and computational approaches in yeast. Here, we review this progress as well as several still outstanding questions.
    DOI:  https://doi.org/10.1016/j.gde.2022.101964
  8. J Biol Chem. 2022 Aug 04. pii: S0021-9258(22)00792-X. [Epub ahead of print] 102349
    An intrinsically disordered transcription activation domain increases the DNA binding affinity and reduces the specificity of NFκB p50/RelA.
    Hannah E R Baughman, Dominic Narang, Wei Chen, Amalia C Villagrán Suárez, Joan Lee, Maxwell J Bachochin, Tristan R Gunther, Peter G Wolynes, Elizabeth A Komives.
      Many transcription factors contain intrinsically disordered transcription activation domains (TADs), which mediate interactions with co-activators to activate transcription. Historically, DNA-binding domains and TADs have been considered as modular units, but recent studies have shown that TADs can influence DNA binding. Whether these results can be generalized to more TADs is not clear. Here we biophysically characterized the NFκB p50/RelA heterodimer including the RelA TAD and investigated the TAD's influence on NFκB-DNA interactions. In solution, we show the RelA TAD is disordered but compact, with helical tendency in two regions that interact with co-activators. We determined that the presence of the TAD increased the stoichiometry of NFκB-DNA complexes containing promoter DNA sequences with tandem κB recognition motifs by promoting the binding of NFκB dimers in excess of the number of κB sites. In addition, we measured the binding affinity of p50/RelA for DNA containing tandem κB sites and single κB sites. While the presence of the TAD enhanced the binding affinity of p50/RelA for all κB sequences tested, it also increased the affinity for non-specific DNA sequences by over 10-fold, leading to an overall decrease in specificity for κB DNA sequences. In contrast, previous studies have generally reported that TADs decrease DNA binding affinity and increase sequence specificity. Our results reveal a novel function of the RelA TAD in promoting binding to non-consensus DNA, which sheds light on previous observations of extensive non-consensus DNA binding by NFκB in vivo in response to strong inflammatory signals.
    Keywords:  DNA-protein interaction; NF‐kB transcription factor; cooperativity; hydrogen-deuterium exchange; intrinsically disordered protein; small-angle X-ray scattering (SAXS); structural model; transcription
    DOI:  https://doi.org/10.1016/j.jbc.2022.102349
  9. Front Plant Sci. 2022 ;13 935819
    TGA transcription factors-Structural characteristics as basis for functional variability.
    Špela Tomaž, Kristina Gruden, Anna Coll.
      TGA transcription factors are essential regulators of various cellular processes, their activity connected to different hormonal pathways, interacting proteins and regulatory elements. Belonging to the basic region leucine zipper (bZIP) family, TGAs operate by binding to their target DNA sequence as dimers through a conserved bZIP domain. Despite sharing the core DNA-binding sequence, the TGA paralogues exert somewhat different DNA-binding preferences. Sequence variability of their N- and C-terminal protein parts indicates their importance in defining TGA functional specificity through interactions with diverse proteins, affecting their DNA-binding properties. In this review, we provide a short and concise summary on plant TGA transcription factors from a structural point of view, including the relation of their structural characteristics to their functional roles in transcription regulation.
    Keywords:  DOG1 domain; TGA transcription factors; functional variability; intrinsically disordered regions; plant transcription regulation; post-translational modifications; structural characteristics
    DOI:  https://doi.org/10.3389/fpls.2022.935819
  10. Phys Chem Chem Phys. 2022 Aug 09.
    Mechanism underlying liquid-to-solid phase transition in fused in sarcoma liquid droplets.
    Shujie Li, Takuya Yoshizawa, Yutaro Shiramasa, Mako Kanamaru, Fumika Ide, Keiji Kitamura, Norika Kashiwagi, Naoya Sasahara, Soichiro Kitazawa, Ryo Kitahara.
      The RNA-binding protein fused in sarcoma (FUS) forms ribonucleoprotein granules via liquid-liquid phase separation (LLPS) in the cytoplasm. The phase separation of FUS accelerates aberrant liquid-solid phase separation and leads to the onset of familial amyotrophic lateral sclerosis (ALS). We previously found that FUS forms two types of liquid condensates in equilibrium, specifically LP-LLPS (i.e., normal type) and HP-LLPS (i.e., aberrant type), each with different partial molar volumes. However, it is unclear how liquid condensates are converted to the pathogenic solid phase. Here, we report a mechanism underlying the aberrant liquid-to-solid phase transition of FUS liquid condensates and the inhibition of this transition with small molecules. We found that the liquid condensate formed via HP-LLPS had greatly reduced dynamics, which is a common feature of aged wild-type FUS droplets and the droplet-like assembly of the ALS patient-type FUS variant. The longer FUS remained on the HP-LLPS, the harder it was to transform it into a mixed state (i.e., one-phase). These results indicate that liquid-to-solid phase transition, namely the aging of droplets, is accelerated with HP-LLPS. Interestingly, arginine suppressed the aging of droplets and HP-LLPS formation more strongly than LP-LLPS formation. These data indicate that the formation of HP-LLPS via the one-phase state or LP-LLPS is a pathway leading to irreversible solid aggregates. Dopamine and pyrocatechol also suppressed HP-LLPS formation. Our data highlight the potential of HP-LLPS to be used as a therapeutic target and arginine as a plausible drug candidate for ALS-causing FUS.
    DOI:  https://doi.org/10.1039/d2cp02171d
  11. Nat Commun. 2022 Aug 06. 13(1): 4586
    Molecular mechanism for the synchronized electrostatic coacervation and co-aggregation of alpha-synuclein and tau.
    Pablo Gracia, David Polanco, Jorge Tarancón-Díez, Ilenia Serra, Maruan Bracci, Javier Oroz, Douglas V Laurents, Inés García, Nunilo Cremades.
      Amyloid aggregation of α-synuclein (αS) is the hallmark of Parkinson's disease and other synucleinopathies. Recently, Tau protein, generally associated with Alzheimer's disease, has been linked to αS pathology and observed to co-localize in αS-rich disease inclusions, although the molecular mechanisms for the co-aggregation of both proteins remain elusive. We report here that αS phase-separates into liquid condensates by electrostatic complex coacervation with positively charged polypeptides such as Tau. Condensates undergo either fast gelation or coalescence followed by slow amyloid aggregation depending on the affinity of αS for the poly-cation and the rate of valence exhaustion of the condensate network. By combining a set of advanced biophysical techniques, we have been able to characterize αS/Tau liquid-liquid phase separation and identified key factors that lead to the formation of hetero-aggregates containing both proteins in the interior of the liquid protein condensates.
    DOI:  https://doi.org/10.1038/s41467-022-32350-9
  12. Front Mol Neurosci. 2022 ;15 964488
    NKCC1 and KCC2: Structural insights into phospho-regulation.
    Anna-Maria Hartmann, Hans Gerd Nothwang.
      Inhibitory neurotransmission plays a fundamental role in the central nervous system, with about 30-50% of synaptic connections being inhibitory. The action of both inhibitory neurotransmitter, gamma-aminobutyric-acid (GABA) and glycine, mainly relies on the intracellular Cl- concentration in neurons. This is set by the interplay of the cation chloride cotransporters NKCC1 (Na+, K+, Cl- cotransporter), a main Cl- uptake transporter, and KCC2 (K+, Cl- cotransporter), the principle Cl- extruder in neurons. Accordingly, their dysfunction is associated with severe neurological, psychiatric, and neurodegenerative disorders. This has triggered great interest in understanding their regulation, with a strong focus on phosphorylation. Recent structural data by cryogenic electron microscopy provide the unique possibility to gain insight into the action of these phosphorylations. Interestingly, in KCC2, six out of ten (60%) known regulatory phospho-sites reside within a region of 134 amino acid residues (12% of the total residues) between helices α8 and α9 that lacks fixed or ordered three-dimensional structures. It thus represents a so-called intrinsically disordered region. Two further phospho-sites, Tyr903 and Thr906, are also located in a disordered region between the ß8 strand and the α8 helix. We make the case that especially the disordered region between helices α8 and α9 acts as a platform to integrate different signaling pathways and simultaneously constitute a flexible, highly dynamic linker that can survey a wide variety of distinct conformations. As each conformation can have distinct binding affinities and specificity properties, this enables regulation of [Cl-] i and thus the ionic driving force in a history-dependent way. This region might thus act as a molecular processor underlying the well described phenomenon of ionic plasticity that has been ascribed to inhibitory neurotransmission. Finally, it might explain the stunning long-range effects of mutations on phospho-sites in KCC2.
    Keywords:  CCC; conformational changes; intrinsically disordered region; neurological diseases; phosphorylation; structure; synaptic inhibition
    DOI:  https://doi.org/10.3389/fnmol.2022.964488
  13. Biochemistry. 2022 Aug 11.
    Phase Separation Properties in Transcriptional Organization.
    Chenghao Guo, Zhuojuan Luo, Chengqi Lin.
      Transcription is of great importance to stress response, fate control, and development, involving the functional cooperation of a large number of transcription factors and cofactors. Transcription machineries assemble rapidly to respond to the physiological and functional needs of cells. Recently, phase-separated biomolecular condensates have emerged as a universal biophysical basis for the spatiotemporal coordination of various cellular activities, including transcription. Here, we summarize and discuss recent advances in understanding of how phase separation contributes to RNA polymerase II (Pol II)-mediated transcriptional regulation, with a focus on the physical properties and dynamics of transcriptional condensates.
    DOI:  https://doi.org/10.1021/acs.biochem.2c00220
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