bims-indpro 2022-06-19 papers

bims-indpro

Biomed News

on Intrinsically disordered proteins

Issue of 2022–06–19
seven papers selected by
Sara Mingu, Johannes Gutenberg University

The intrinsically disordered region from PP2C phosphatases functions as a conserved CO2 sensor.
Conformational ensemble of the full-length SARS-CoV-2 nucleocapsid (N) protein based on molecular simulations and SAXS data.
A little less aggregation a little more replication: Viral manipulation of stress granules.
Wide-Line NMR Melting Diagrams, Their Thermodynamic Interpretation, and Secondary Structure Predictions for A30P and E46K α-Synuclein.
Hidden multivalency in phosphatase recruitment by a disordered AKAP scaffold.
Backbone resonance assignments and dynamics of S. cerevisiae SERF.
Insights into the Atomistic Mechanisms of Phosphorylation in Disrupting Liquid-Liquid Phase Separation and Aggregation of the FUS Low-Complexity Domain.

Nat Cell Biol. 2022 Jun 16.

The intrinsically disordered region from PP2C phosphatases functions as a conserved CO2 sensor.

Mao Zhang, Cheng Zhu, Yuanyuan Duan, Tongbao Liu, Haoping Liu, Chang Su, Yang Lu.

Carbon dioxide not only plays a central role in the carbon cycle, but also acts as a crucial signal in living cells. Adaptation to changing CO2 concentrations is critical for all organisms. Conversion of CO2 to HCO3- by carbonic anhydrase and subsequent HCO3--triggered signalling are thought to be important for cellular responses to CO2 (refs. 1-3). However, carbonic anhydrases are suggested to transduce a change in CO2 rather than be a direct CO2 sensor4,5, the mechanism(s) by which organisms sense CO2 remain unknown. Here we demonstrate that a unique group of PP2C phosphatases from fungi and plants senses CO2, but not HCO3-, to control diverse cellular programmes. Different from other phosphatases, these PP2Cs all have an intrinsically disordered region (IDR). They formed reversible liquid-like droplets through phase separation both in cells and in vitro, and were activated in response to elevated environmental CO2 in an IDR-dependent manner. The IDRs in PP2Cs are characterized by a sequence of polar amino acids enriched in serine/threonine, which provides CO2 responsiveness. CO2-responsive activation of PP2Cs via the serine/threonine-rich IDR-mediated phase separation represents a direct CO2 sensing mechanism and is widely exploited.

DOI: https://doi.org/10.1038/s41556-022-00936-6
Biophys Chem. 2022 Jun 07. pii: S0301-4622(22)00085-0. [Epub ahead of print]288 106843

Conformational ensemble of the full-length SARS-CoV-2 nucleocapsid (N) protein based on molecular simulations and SAXS data.

Bartosz Różycki, Evzen Boura.

  The nucleocapsid protein of the SARS-CoV-2 virus comprises two RNA-binding domains and three regions that are intrinsically disordered. While the structures of the RNA-binding domains have been solved using protein crystallography and NMR, current knowledge of the conformations of the full-length nucleocapsid protein is rather limited. To fill in this knowledge gap, we combined coarse-grained molecular simulations with data from small-angle X-ray scattering (SAXS) experiments using the ensemble refinement of SAXS (EROS) method. Our results show that the dimer of the full-length nucleocapsid protein exhibits large conformational fluctuations with its radius of gyration ranging from about 4 to 8 nm. The RNA-binding domains do not make direct contacts. The disordered region that links these two domains comprises a hydrophobic α-helix which makes frequent and nonspecific contacts with the RNA-binding domains. Each of the intrinsically disordered regions adopts conformations that are locally compact, yet on average, much more extended than Gaussian chains of equivalent lengths. We offer a detailed picture of the conformational ensemble of the nucleocapsid protein dimer under near-physiological conditions, which will be important for understanding the nucleocapsid assembly process.

Keywords:  EROS; Nucleocapsid; SARS-CoV-2; SAXS

DOI:  https://doi.org/10.1016/j.bpc.2022.106843
Wiley Interdiscip Rev RNA. 2022 Jun 16. e1741

A little less aggregation a little more replication: Viral manipulation of stress granules.

Matthew J Brownsword, Nicolas Locker.

  Recent exciting studies have uncovered how membrane-less organelles, also known as biocondensates, are providing cells with rapid response pathways, allowing them to re-organize their cellular contents and adapt to stressful conditions. Their assembly is driven by the phase separation of their RNAs and intrinsically disordered protein components into condensed foci. Among these, stress granules (SGs) are dynamic cytoplasmic biocondensates that form in response to many stresses, including activation of the integrated stress response or viral infections. SGs sit at the crossroads between antiviral signaling and translation because they concentrate signaling proteins and components of the innate immune response, in addition to translation machinery and stalled mRNAs. Consequently, they have been proposed to contribute to antiviral activities, and therefore are targeted by viral countermeasures. Equally, SGs components can be commandeered by viruses for their own efficient replication. Phase separation processes are an important component of the viral life cycle, for example, driving the assembly of replication factories or inclusion bodies. Therefore, in this review, we will outline the recent understanding of this complex interplay and tug of war between viruses, SGs, and their components. This article is categorized under: RNA in Disease and Development > RNA in Disease Translation > Regulation RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes.

Keywords:  G3BP1; phase separation; stress granules; stress response; virus

DOI:  https://doi.org/10.1002/wrna.1741
ACS Omega. 2022 Jun 07. 7(22): 18323-18330

Wide-Line NMR Melting Diagrams, Their Thermodynamic Interpretation, and Secondary Structure Predictions for A30P and E46K α-Synuclein.

Mónika Bokor, Eszter Házy, Ágnes Tantos.

Parkinson's disease is thought to be caused by aggregation of the intrinsically disordered protein, α-synuclein. Two amyloidogenic variants, A30P, and E46K familial mutants were investigated by wide-line 1H NMR spectrometry as a completion of our earlier work on wild-type and A53T α-synuclein (Bokor M. et al. WT and A53T α-synuclein systems: melting diagram and its new interpretation. Int. J. Mol. Sci.2020, 21, 3997.). A monolayer of mobile water molecules hydrates A30P α-synuclein at the lowest potential barriers (temperatures), while E46K α-synuclein has here third as much mobile hydration, insufficient for functionality. According to wide-line 1H NMR results and secondary structure predictions, E46K α-synuclein is more compact than the A30P variant and they are more compact than the wild type (WT) and A53T variants. Linear hydration vs potential barrier sections of A30P α-synuclein shows one and E46K shows two slopes. The different slopes of the latter between potential barriers E a,1 and E a,2 reflect a change in water-protein interactions. The 31-32% homogeneous potential barrier distribution of the protein-water bonds refers to a non-negligible amount of secondary structures in all four α-synuclein variants. The secondary structures detected by wide-line 1H NMR are solvent-exposed α-helices, which are predicted by secondary structure models. β-sheets are only minor components of the protein structures as three- and eight-state predicted secondary structures are dominated by α-helices and coils.

DOI: https://doi.org/10.1021/acsomega.2c00477
J Mol Biol. 2022 Jun 10. pii: S0022-2836(22)00274-1. [Epub ahead of print] 167682

Hidden multivalency in phosphatase recruitment by a disordered AKAP scaffold.

Matthew Watson, Teresa B Almeida, Arundhati Ray, Christina Hanack, Rory Elston, Joan Btesh, Peter A McNaughton, Katherine Stott.

  Disordered scaffold proteins provide multivalent landing pads that, via a series of embedded Short Linear Motifs (SLiMs), bring together the components of a complex to orchestrate precise spatial and temporal regulation of cellular processes. One such protein is AKAP5 (previously AKAP79), which contains SLiMs that anchor PKA and Calcineurin, and recruit substrate (the TRPV1 receptor). Calcineurin is anchored to AKAP5 by a well-characterised PxIxIT SLiM. Here we show, using a combination of biochemical and biophysical approaches, that the Calcineurin PxIxIT-binding groove also recognises several hitherto unknown lower-affinity SLiMs in addition to the PxIxIT motif. We demonstrate that the assembly is in reality a complex system with conserved SLiMs spanning a wide affinity range. The capture is analogous to that seen for many DNA-binding proteins that have a weak non-specific affinity for DNA outside the canonical binding site, but different in that it involves (i) two proteins, and (ii) hydrophobic rather than electrostatic interactions. It is also compatible with the requirement for both stable anchoring of the enzyme and responsive downstream signalling. We conclude that the AKAP5 C-terminus is enriched in lower-affinity/mini-SLiMs that, together with the canonical SLiM, maintain a structurally disordered but tightly regulated signalosome.

Keywords:  A-kinase anchoring protein (AKAP); Calcineurin; allovalency; intrinsically disordered protein; multivalency; short linear motif (SLiM)

DOI:  https://doi.org/10.1016/j.jmb.2022.167682
Biomol NMR Assign. 2022 Jun 17.

Backbone resonance assignments and dynamics of S. cerevisiae SERF.

Yicong Liu, Chaozhe Wang, Yangzhuoyue Jin, Guosheng Jiang, Lichun He, Maili Liu.

  Abnormal protein aggregation and precipitation are associated with the perturbation of cellular function and underlie a variety of neurodegenerative diseases. S. cerevisiae SERF (ScSERF), a homolog of modifier of aggregation-4 (MOAG-4) and small EDRK-rich factor protein (SERF1a) is highly conserved and discovered as an enhancer of amyloid formation of Aβ40 and α-synuclein both in vitro and in vivo. However, the detailed molecular mechanism whereby ScSERF and its homologs accelerate amyloid formation is not well known yet. Herein, we present the 1 H, 15 N and 13 C NMR assignments of the 68 amino acids long ScSERF. Although ScSERF displays a very high degree of disorder, secondary chemical shifts of Cα, Cβ, 15 N{1 H}-NOE values and the residue-specific secondary structure propensity (SSP) scores indicate the segment spanning residues 36E-65 K has a strong helical propensity. This work sets the stage for further detailed structural and dynamic investigations of ScSERF and the molecular mechanism it utilizes in accelerating amyloid formation.

Keywords:  Assignments; Intrinsically disordered protein; NMR; ScSERF

DOI:  https://doi.org/10.1007/s12104-022-10077-4
J Chem Inf Model. 2022 Jun 16.

Insights into the Atomistic Mechanisms of Phosphorylation in Disrupting Liquid-Liquid Phase Separation and Aggregation of the FUS Low-Complexity Domain.

Zenghui Lao, Xuewei Dong, Xianshi Liu, Fangying Li, Yujie Chen, Yiming Tang, Guanghong Wei.

Fused in sarcoma (FUS), a nuclear RNA binding protein, can not only undergo liquid-liquid phase separation (LLPS) to form dynamic biomolecular condensates but also aggregate into solid amyloid fibrils which are associated with the pathology of amyotrophic lateral sclerosis and frontotemporal lobar degeneration diseases. Phosphorylation in the FUS low-complexity domain (FUS-LC) inhibits FUS LLPS and aggregation. However, it remains largely elusive what are the underlying atomistic mechanisms of this inhibitory effect and whether phosphorylation can disrupt preformed FUS fibrils, reversing the FUS gel/solid phase toward the liquid phase. Herein, we systematically investigate the impacts of phosphorylation on the conformational ensemble of the FUS37-97 monomer and dimer and the structure of the FUS37-97 fibril by performing extensive all-atom molecular dynamics simulations. Our simulations reveal three key findings: (1) phosphorylation shifts the conformations of FUS37-97 from the β-rich, fibril-competent state toward a helix-rich, fibril-incompetent state; (2) phosphorylation significantly weakens protein-protein interactions and enhances protein-water interactions, which disfavor FUS-LC LLPS as well as aggregation and facilitate the dissolution of the preformed FUS-LC fibril; and (3) the FUS37-97 peptide displays a high β-strand probability in the region spanning residues 52-67, and phosphorylation at S54 and S61 residues located in this region is crucial for the disruption of LLPS and aggregation of FUS-LC. This study may pave the way for ameliorating phase-separation-related pathologies via site-specific phosphorylation.

DOI: https://doi.org/10.1021/acs.jcim.2c00414