bims-indpro 2022-10-30 papers

bims-indpro

Biomed News

on Intrinsically disordered proteins

Issue of 2022–10–30
24 papers selected by
Sara Mingu, Johannes Gutenberg University

Deciphering the Alphabet of Disorder-Glu and Asp Act Differently on Local but Not Global Properties.
Biophysical insights into glucose-dependent transcriptional regulation by PDX1.
Single-chain and condensed-state behavior of hnRNPA1 from molecular simulations.
Looking at the Pathogenesis of the Rabies Lyssavirus Strain Pasteur Vaccins through a Prism of the Disorder-Based Bioinformatics.
Small molecules targeting the disordered transactivation domain of the androgen receptor induce the formation of collapsed helical states.
The chaperone DNAJB6 surveils FG-nucleoporins and is required for interphase nuclear pore complex biogenesis.
Intrinsically disordered plant protein PARCL co-localizes with RNA in phase-separated condensates whose formation can be regulated by mutating the PLD.
Hotspot mutations in the structured ENL YEATS domain link aberrant transcriptional condensates and cancer.
PABPN1 functions as a hub in the assembly of nuclear poly(A) domains that are essential for mouse oocyte development.
Study on internal structure of casein micelles in reconstituted skim milk powder.
TransDFL: Identification of Disordered Flexible Linkers in Proteins by Transfer Learning.
Molecular Insights into the Misfolding and Dimerization Dynamics of the Full-Length α-Synuclein from Atomistic Discrete Molecular Dynamics Simulations.
The C-Terminus of the PSMA3 Proteasome Subunit Preferentially Traps Intrinsically Disordered Proteins for Degradation.
The Epigenetic Dimension of Protein Structure Is an Intrinsic Weakness of the AlphaFold Program.
Beyond Microsatellite Instability: Intrinsic Disorder as a Potential Link Between Protein Short Tandem Repeats and Cancer.
Nanomedicines Meet Disordered Proteins: A Shift from Traditional Materials and Concepts to Innovative Polymers.
The Effect of Positive Charge Distribution on the Cryoprotective Activity of Dehydrins.
Intrinsic Disorder in BAP1 and Its Association with Uveal Melanoma.
Intrinsic protein disorder uncouples affinity from binding specificity.
SETH predicts nuances of residue disorder from protein embeddings.
Does one plus one always equal two? Structural differences between nesfatin-1, -2, and nesfatin-1/2.
Functional Tuning of Intrinsically Disordered Regions in Human Proteins by Composition Bias.
A sequence-based foldability score combined with AlphaFold2 predictions to disentangle the protein order/disorder continuum.
Reorganization of Cell Compartmentalization Induced by Stress.

Biomolecules. 2022 Oct 04. pii: 1426. [Epub ahead of print]12(10):

Deciphering the Alphabet of Disorder-Glu and Asp Act Differently on Local but Not Global Properties.

Mette Ahrensback Roesgaard, Jeppe E Lundsgaard, Estella A Newcombe, Nina L Jacobsen, Francesco Pesce, Emil E Tranchant, Søren Lindemose, Andreas Prestel, Rasmus Hartmann-Petersen, Kresten Lindorff-Larsen, Birthe B Kragelund.

  Compared to folded proteins, the sequences of intrinsically disordered proteins (IDPs) are enriched in polar and charged amino acids. Glutamate is one of the most enriched amino acids in IDPs, while the chemically similar amino acid aspartate is less enriched. So far, the underlying functional differences between glutamates and aspartates in IDPs remain poorly understood. In this study, we examine the differential effects of aspartate and glutamates in IDPs by comparing the function and conformational ensemble of glutamate and aspartate variants of the disordered protein Dss1, using a range of assays, including interaction studies, nuclear magnetic resonance spectroscopy, small-angle X-ray scattering and molecular dynamics simulation. First, we analyze the sequences of the rapidly growing database of experimentally verified IDPs (DisProt) and show that glutamate enrichment is not caused by a taxonomy bias in IDPs. From analyses of local and global structural properties as well as cell growth and protein-protein interactions using a model acidic IDP from yeast and three Glu/Asp variants, we find that while the Glu/Asp variants support similar function and global dimensions, the variants differ in their binding affinities and population of local transient structural elements. We speculate that these local structural differences may play roles in functional diversity, where glutamates can support increased helicity, important for folding and binding, while aspartates support extended structures and form helical caps, as well as playing more relevant roles in, e.g., transactivation domains and ion-binding.

Keywords:  Dss1; IDPs; NMR; SAXS; intrinsically disordered protein; molecular dynamics; sequence composition

DOI:  https://doi.org/10.3390/biom12101426
J Biol Chem. 2022 Oct 19. pii: S0021-9258(22)01066-3. [Epub ahead of print] 102623

Biophysical insights into glucose-dependent transcriptional regulation by PDX1.

Emery T Usher, Scott A Showalter.

  The pancreatic and duodenal homeobox 1 (PDX1) is a central regulator of glucose-dependent transcription of insulin in pancreatic β cells. PDX1 transcription factor activity is integral to the development and sustained health of the pancreas; accordingly, deciphering the complex network of cellular cues that lead to PDX1 activation or inactivation is an important step toward understanding the etiopathologies of pancreatic diseases and the development of novel therapeutics. Despite nearly three decades of research into PDX1 control of Insulin expression, the molecular mechanisms that dictate the function of PDX1 in response to glucose are still elusive. The transcriptional activation functions of PDX1 are regulated, in part, by its two intrinsically disordered regions (IDRs) which pose a barrier to its structural and biophysical characterization. Indeed, many studies of PDX1 interactions, clinical mutations, and post-translational modifications lack molecular-level detail. Emerging methods for the quantitative study of IDRs and refined models for transactivation now enable us to validate and interrogate the biochemical and biophysical features of PDX1 that dictate its function. The goal of this review is to summarize existing PDX1 studies and, further, to generate a comprehensive resource for future studies of transcriptional control via PDX1.

Keywords:  biophysics; diabetes; gene regulation; intrinsically disordered protein; pancreatic and duodenal homeobox 1; post-translational modification; transcription factor

DOI:  https://doi.org/10.1016/j.jbc.2022.102623
J Chem Phys. 2022 Oct 21. 157(15): 154903

Single-chain and condensed-state behavior of hnRNPA1 from molecular simulations.

D Janka Bauer, Lukas S Stelzl, Arash Nikoubashman.

Intrinsically disordered proteins (IDPs) are essential components for the formation of membraneless organelles, which play key functional and regulatory roles within biological systems. These complex assemblies form and dissolve spontaneously over time via liquid-liquid phase separation of IDPs. Mutations in their amino acid sequence can alter their phase behavior, which has been linked to the emergence of severe diseases. We study the conformation and phase behavior of a low-complexity domain of heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1) using coarse-grained implicit solvent molecular dynamics simulations. We systematically analyze how these properties are affected by the number of aromatic residues within the examined sequences. We find a significant compaction of the chains and an increase in the critical temperature with an increasing number of aromatic residues. The local persistence length is determined in single-chain simulations, revealing strong sequence-specific variations along the chain contour. Comparing single-chain and condensed-state simulations, we find many more collapsed polymer conformations in the dilute systems, even at temperatures near the estimated θ-temperature of the solution. These observations strongly support the hypothesis that aromatic residues play a dominant role in condensation, which is further corroborated by a detailed analysis of the intermolecular contacts, and conversely that important properties of condensates are captured in coarse-grained simulations. Interestingly, we observe density inhomogeneities within the condensates near criticality, which are driven by electrostatic interactions. Finally, we find that the relatively small fraction of hydrophobic residues in the IDPs results in interfacial tensions, which are significantly lower compared to typical combinations of immiscible simple liquids.

DOI: https://doi.org/10.1063/5.0105540
Biomolecules. 2022 Oct 07. pii: 1436. [Epub ahead of print]12(10):

Looking at the Pathogenesis of the Rabies Lyssavirus Strain Pasteur Vaccins through a Prism of the Disorder-Based Bioinformatics.

Surya Dhulipala, Vladimir N Uversky.

  Rabies is a neurological disease that causes between 40,000 and 70,000 deaths every year. Once a rabies patient has become symptomatic, there is no effective treatment for the illness, and in unvaccinated individuals, the case-fatality rate of rabies is close to 100%. French scientists Louis Pasteur and Émile Roux developed the first vaccine for rabies in 1885. If administered before the virus reaches the brain, the modern rabies vaccine imparts long-lasting immunity to the virus and saves more than 250,000 people every year. However, the rabies virus can suppress the host's immune response once it has entered the cells of the brain, making death likely. This study aimed to make use of disorder-based proteomics and bioinformatics to determine the potential impact that intrinsically disordered protein regions (IDPRs) in the proteome of the rabies virus might have on the infectivity and lethality of the disease. This study used the proteome of the Rabies lyssavirus (RABV) strain Pasteur Vaccins (PV), one of the best-understood strains due to its use in the first rabies vaccine, as a model. The data reported in this study are in line with the hypothesis that high levels of intrinsic disorder in the phosphoprotein (P-protein) and nucleoprotein (N-protein) allow them to participate in the creation of Negri bodies and might help this virus to suppress the antiviral immune response in the host cells. Additionally, the study suggests that there could be a link between disorder in the matrix (M) protein and the modulation of viral transcription. The disordered regions in the M-protein might have a possible role in initiating viral budding within the cell. Furthermore, we checked the prevalence of functional disorder in a set of 37 host proteins directly involved in the interaction with the RABV proteins. The hope is that these new insights will aid in the development of treatments for rabies that are effective after infection.

Keywords:  intrinsic disorder; intrinsically disordered protein; intrinsically disordered protein region; protein–protein interaction; rabies

DOI:  https://doi.org/10.3390/biom12101436
Nat Commun. 2022 Oct 27. 13(1): 6390

Small molecules targeting the disordered transactivation domain of the androgen receptor induce the formation of collapsed helical states.

Jiaqi Zhu, Xavier Salvatella, Paul Robustelli.

Intrinsically disordered proteins, which do not adopt well-defined structures under physiological conditions, are implicated in many human diseases. Small molecules that target the disordered transactivation domain of the androgen receptor have entered human trials for the treatment of castration-resistant prostate cancer (CRPC), but no structural or mechanistic rationale exists to explain their inhibition mechanisms or relative potencies. Here, we utilize all-atom molecular dynamics computer simulations to elucidate atomically detailed binding mechanisms of the compounds EPI-002 and EPI-7170 to the androgen receptor. Our simulations reveal that both compounds bind at the interface of two transiently helical regions and induce the formation of partially folded collapsed helical states. We find that EPI-7170 binds androgen receptor more tightly than EPI-002 and we identify a network of intermolecular interactions that drives higher affinity binding. Our results suggest strategies for developing more potent androgen receptor inhibitors and general strategies for disordered protein drug design.

DOI: https://doi.org/10.1038/s41467-022-34077-z
Nat Cell Biol. 2022 Oct 27.

The chaperone DNAJB6 surveils FG-nucleoporins and is required for interphase nuclear pore complex biogenesis.

E F Elsiena Kuiper, Paola Gallardo, Tessa Bergsma, Muriel Mari, Maiara Kolbe Musskopf, Jeroen Kuipers, Ben N G Giepmans, Anton Steen, Harm H Kampinga, Liesbeth M Veenhoff, Steven Bergink.

Biogenesis of nuclear pore complexes (NPCs) includes the formation of the permeability barrier composed of phenylalanine-glycine-rich nucleoporins (FG-Nups) that regulate the selective passage of biomolecules across the nuclear envelope. The FG-Nups are intrinsically disordered and prone to liquid-liquid phase separation and aggregation when isolated. How FG-Nups are protected from making inappropriate interactions during NPC biogenesis is not fully understood. Here we find that DNAJB6, a molecular chaperone of the heat shock protein network, forms foci in close proximity to NPCs. The number of these foci decreases upon removal of proteins involved in the early steps of interphase NPC biogenesis. Conversely, when this process is stalled in the last steps, the number of DNAJB6-containing foci increases and these foci are identified as herniations at the nuclear envelope. Immunoelectron tomography shows that DNAJB6 localizes inside the lumen of the herniations arising at NPC biogenesis intermediates. Loss of DNAJB6 results in the accumulation of cytosolic annulate lamellae, which are structures containing partly assembled NPCs, a feature associated with disturbances in NPC biogenesis. We find that DNAJB6 binds to FG-Nups and can prevent the aggregation of the FG region of several FG-Nups in cells and in vitro. Together, our data show that the molecular chaperone DNAJB6 provides quality control during NPC biogenesis and is involved in the surveillance of native intrinsically disordered FG-Nups.

DOI: https://doi.org/10.1038/s41556-022-01010-x
J Biol Chem. 2022 Oct 20. pii: S0021-9258(22)01074-2. [Epub ahead of print] 102631

Intrinsically disordered plant protein PARCL co-localizes with RNA in phase-separated condensates whose formation can be regulated by mutating the PLD.

Anna Ostendorp, Steffen Ostendorp, Yuan Zhou, Zoé Chaudron, Lukas Wolffram, Khadija Rombi, Linn von Pein, Sven Falke, Cy M Jeffries, Dmitri I Svergun, Christian Betzel, Richard J Morris, Friedrich Kragler, Julia Kehr.

  In higher plants, long-distance RNA transport via the phloem is crucial for communication between distant plant tissues to align development with stress responses and reproduction. Several recent studies suggest that specific RNAs are among the potential long-distance information transmitters. However, it is yet not well understood how these RNAs enter the phloem stream, how they are transported, and how they are released at their destination. It was proposed that phloem RNA-binding proteins (RBPs) facilitate RNA translocation. In the present study, we characterized two orthologs of the phloem-associated RNA chaperone-like (PARCL) protein from Arabidopsis thaliana and Brassica napus at functional and structural levels. Microscale thermophoresis (MST) showed that these phloem-abundant proteins can bind a broad spectrum of RNAs and show RNA chaperone activity in FRET-based in vitro assays. Our SAXS experiments revealed a high degree of disorder, typical for RNA-binding proteins. In agroinfiltrated tobacco plants, eYFP-PARCL proteins mainly accumulated in nuclei and nucleoli and formed cytosolic and nuclear condensates. We found that formation of these condensates was impaired by tyrosine-to-glutamate mutations in the predicted prion-like domain (PLD), while C-terminal serine-to-glutamate mutations did not affect condensation but reduced RNA binding and chaperone activity. Furthermore, our in vitro experiments confirmed phase separation of PARCL and co-localization of RNA with the condensates, while mutation as well as phosphorylation of the PLD reduced phase separation. Together, our results suggest that RNA binding and condensate formation of PARCL can be regulated independently by modification of the C-terminus and/ or the PLD.

Keywords:  RNA binding protein; RNP complex; SAXS; intrinsically disordered protein; liquid condensates; microscale thermophoresis; phase separation; phloem; phosphorylation; prion-like domain

DOI:  https://doi.org/10.1016/j.jbc.2022.102631
Mol Cell. 2022 Oct 18. pii: S1097-2765(22)00955-8. [Epub ahead of print]

Hotspot mutations in the structured ENL YEATS domain link aberrant transcriptional condensates and cancer.

Lele Song, Xinyi Yao, Hangpeng Li, Bo Peng, Alan P Boka, Yiman Liu, Guochao Chen, Zhenyang Liu, Kaeli M Mathias, Lingbo Xia, Qinglan Li, Mustafa Mir, Yuanyuan Li, Haitao Li, Liling Wan.

  Growing evidence suggests prevalence of transcriptional condensates on chromatin, yet their mechanisms of formation and functional significance remain largely unclear. In human cancer, a series of mutations in the histone acetylation reader ENL create gain-of-function mutants with increased transcriptional activation ability. Here, we show that these mutations, clustered in ENL's structured acetyl-reading YEATS domain, trigger aberrant condensates at native genomic targets through multivalent homotypic and heterotypic interactions. Mechanistically, mutation-induced structural changes in the YEATS domain, ENL's two disordered regions of opposing charges, and the incorporation of extrinsic elongation factors are all required for ENL condensate formation. Extensive mutagenesis establishes condensate formation as a driver of oncogenic gene activation. Furthermore, expression of ENL mutants beyond the endogenous level leads to non-functional condensates. Our findings provide new mechanistic and functional insights into cancer-associated condensates and support condensate dysregulation as an oncogenic mechanism.

Keywords:  ENL YEATS; IDR; cancer epigenetics; chromatin reader; intrinsically disordered protein region; multivalent interactions; mutations; phase separation; structured domains; transcription elongation; transcriptional condensates/hubs

DOI:  https://doi.org/10.1016/j.molcel.2022.09.034
Sci Adv. 2022 Oct 28. 8(43): eabn9016

PABPN1 functions as a hub in the assembly of nuclear poly(A) domains that are essential for mouse oocyte development.

Xing-Xing Dai, Shuai-Bo Pi, Long-Wen Zhao, Yun-Wen Wu, Jing-Ling Shen, Song-Ying Zhang, Qian-Qian Sha, Heng-Yu Fan.

Growing oocytes store a large amount of maternal mRNA to support the subsequent "maternal-zygotic transition" process. At present, it is not clear how the growing oocytes store and process the newly transcribed mRNA under physiological conditions. In this study, we report non-membrane-bound compartments, nuclear poly(A) domains (NPADs), as the hub for newly transcribed mRNA, in developing mouse oocytes. The RNA binding protein PABPN1 promotes the formation of NPAD through its N-terminal disordered domain and RNA-recognized motif by means of liquid phase separation. Pabpn1-null growing oocytes cannot form NPAD normally in vivo and have defects in stability of oocyte growing-related transcripts and formation of long 3' untranslated region isoform transcripts. Ultimately, Pabpn1fl/fl;Gdf9-Cre mice are completely sterile with primary ovarian insufficiency. These results demonstrate that NPAD formed by the phase separation properties of PABPN1-mRNA are the hub of the newly transcribed mRNA and essential for the development of oocytes and female reproduction.

DOI: https://doi.org/10.1126/sciadv.abn9016
Int J Biol Macromol. 2022 Oct 21. pii: S0141-8130(22)02377-7. [Epub ahead of print]

Study on internal structure of casein micelles in reconstituted skim milk powder.

Zhongyao Du, Ning Xu, Ying Yang, Guimei Li, Zhonghong Tai, Na Li, Yang Sun.

  In the current study, the internal structure of casein micelles (CMs), primary casein cluster, has been studied by size-exclusion chromatography (SEC) coupled with small-angle X-ray scattering (SAXS), isothermal titration calorimetry (ITC), transmission electron microscopy (TEM) and molecular dynamics (MD) simulations. The casein cluster featured a hierarchical structure predominately consisting of αs-CN and β-CN molecules and a trace of κ-CN. ITC profile showed a typical enthalpogram of CMs with a critical micelle concentration (CMC) of ~0.85 μg/mL. The casein cluster exhibited apparent characteristics of intrinsically disordered proteins (IDPs) with a secondary structure content of 24 % in α-helix, 35.4 % in antiparallel-parallel, 20.2 % in β-turn and 20.4 % in random coil. SAXS results revealed a slightly elongated and tortuous worm-like conformation for the casein cluster in solution with an aspect ratio of 1.36 and an estimated molecular weight of 162.7 kDa. Further scattering data analysis proposed three coexisted species (αs1-β-αs2-CN, αs1-CN and αs1-β-αs2-CN dimer) with a volume fraction of 57.4 %, 30.1 %, and 12.5 % respectively in the casein cluster. TEM observation was consistent with the results of spectra, SAXS and MD that calcium sequestration resulted in a more extended and loose structure, instead, EDTA chelation induced a more compact conformation of the casein cluster.

Keywords:  Casein micelles; Integrative structural biology; Internal structure; Solution SAXS

DOI:  https://doi.org/10.1016/j.ijbiomac.2022.10.135
Genomics Proteomics Bioinformatics. 2022 Oct 19. pii: S1672-0229(22)00132-2. [Epub ahead of print]

TransDFL: Identification of Disordered Flexible Linkers in Proteins by Transfer Learning.

Yihe Pang, Bin Liu.

  Disordered flexible linkers (DFLs) are the functional disordered regions in proteins, which are the sub-regions of intrinsically disordered regions (IDRs) and play important roles in connecting domains and maintaining inter-domain interactions. Trained with the limited available DFLs, the existing DFL predictors based on the machine learning techniques tend to predict the ordered residues as DFLs leading to a high false-positive rate (FPR) and low prediction accuracy. Previous studies have shown that DFLs are extremely flexible disordered regions, which are usually predicted as disordered residues with high confidence [P(D) > 0.9] by an IDR predictor. Therefore, transferring an IDR predictor to an accurate DFL predictor is of great significance for understanding the functions of IDRs. In this study, we proposed a new predictor called TransDFL for identifying DFLs by transferring the RFPR-IDP predictor for IDR identification to the DFL prediction. The RFPR-IDP was pre-trained with IDR sequences to learn the general features between IDRs and DFLs, which is helpful to reduce the false positives in the ordered regions. RFPR-IDP was fine-tuned with the DFL sequences to capture the specific features of DFLs so as to be transferred into the TransDFL. Experimental results of two application scenarios (prediction of DFLs only in the IDRs or prediction of DFLs in the entire proteins) showed that TransDFL consistently outperforms the other existing DFL predictors with higher accuracy. The corresponding web server of TransDFL can be freely accessed from http://bliulab.net/TransDFL/.

Keywords:  Computational predictor; Disordered flexible linkers; False-positive rate; Intrinsically disordered proteins; Transfer learning

DOI:  https://doi.org/10.1016/j.gpb.2022.10.004
ACS Chem Neurosci. 2022 Oct 24.

Molecular Insights into the Misfolding and Dimerization Dynamics of the Full-Length α-Synuclein from Atomistic Discrete Molecular Dynamics Simulations.

Yu Zhang, Ying Wang, Yuying Liu, Guanghong Wei, Feng Ding, Yunxiang Sun.

  The misfolding and pathological aggregation of α-synuclein forming insoluble amyloid deposits is associated with Parkinson's disease, the second most common neurodegenerative disease in the world population. Characterizing the self-assembly mechanism of α-synuclein is critical for discovering treatments against synucleinopathies. The intrinsically disordered property, high degrees of freedom, and macroscopic timescales of conformational conversion make its characterization extremely challenging in vitro and in silico. Here, we systematically investigated the dynamics of monomer misfolding and dimerization of the full-length α-synuclein using atomistic discrete molecular dynamics simulations. Our results suggested that both α-synuclein monomers and dimers mainly adopted unstructured formations with partial helices around the N-terminus (residues 8-32) and various β-sheets spanning the residues 35-56 (N-terminal tail) and residues 61-95 (NAC region). The C-terminus mostly assumed an unstructured formation wrapping around the lateral surface and the elongation edge of the β-sheet core formed by an N-terminal tail and NAC regions. Dimerization enhanced the β-sheet formation along with a decrease in the unstructured content. The inter-peptide β-sheets were mainly formed by the N-terminal tail and NACore (residues 68-78) regions, suggesting that these two regions played critical roles in the amyloid aggregation of α-synuclein. Interactions of the C-terminus with the N-terminal tail and the NAC region were significantly suppressed in the α-synuclein dimer, indicating that the interaction of the C-terminus with the N-terminal tail and NAC regions could prevent α-synuclein aggregation. These results on the structural ensembles and early aggregation dynamics of α-synuclein will help understand the nucleation and fibrillization of α-synuclein.

Keywords:  DMD simulation; Parkinson’s disease; amyloid; dimerization; α-synuclein

DOI:  https://doi.org/10.1021/acschemneuro.2c00531
Cells. 2022 Oct 14. pii: 3231. [Epub ahead of print]11(20):

The C-Terminus of the PSMA3 Proteasome Subunit Preferentially Traps Intrinsically Disordered Proteins for Degradation.

Assaf Biran, Nadav Myers, Shirel Steinberger, Julia Adler, Marianna Riutin, Karin Broennimann, Nina Reuven, Yosef Shaul.

  The degradation of intrinsically disordered proteins (IDPs) by a non-26S proteasome process does not require proteasomal targeting by polyubiquitin. However, whether and how IDPs are recognized by the non-26S proteasome, including the 20S complex, remains unknown. Analyses of protein interactome datasets revealed that the 20S proteasome subunit, PSMA3, preferentially interacts with many IDPs. In vivo and cell-free experiments revealed that the C-terminus of PSMA3, a 69-amino-acids-long fragment, is an IDP trapper. A recombinant trapper is sufficient to interact with many IDPs, and blocks IDP degradation in vitro by the 20S proteasome, possibly by competing with the native trapper. In addition, over a third of the PSMA3 trapper-binding proteins have previously been identified as 20S proteasome substrates and, based on published datasets, many of the trapper-binding proteins are associated with the intracellular proteasomes. The PSMA3-trapped IDPs that are proteasome substrates have the unique features previously recognized as characteristic 20S proteasome substrates in vitro. We propose a model whereby the PSMA3 C-terminal region traps a subset of IDPs to facilitate their proteasomal degradation.

Keywords:  20S proteasome; intrinsically disordered proteins; proteasomal degradation; proteostasis

DOI:  https://doi.org/10.3390/cells11203231
Biomolecules. 2022 Oct 20. pii: 1527. [Epub ahead of print]12(10):

The Epigenetic Dimension of Protein Structure Is an Intrinsic Weakness of the AlphaFold Program.

Fodil Azzaz, Nouara Yahi, Henri Chahinian, Jacques Fantini.

  One of the most important lessons we have learned from sequencing the human genome is that not all proteins have a 3D structure. In fact, a large part of the human proteome is made up of intrinsically disordered proteins (IDPs) which can adopt multiple structures, and therefore, multiple functions, depending on the ligands with which they interact. Under these conditions, one can wonder about the value of algorithms developed for predicting the structure of proteins, in particular AlphaFold, an AI which claims to have solved the problem of protein structure. In a recent study, we highlighted a particular weakness of AlphaFold for membrane proteins. Based on this observation, we have proposed a paradigm, referred to as "Epigenetic Dimension of Protein Structure" (EDPS), which takes into account all environmental parameters that control the structure of a protein beyond the amino acid sequence (hence "epigenetic"). In this new study, we compare the reliability of the AlphaFold and Robetta algorithms' predictions for a new set of membrane proteins involved in human pathologies. We found that Robetta was generally more accurate than AlphaFold for ascribing a membrane-compatible topology. Raft lipids (e.g., gangliosides), which control the structural dynamics of membrane protein structure through chaperone effects, were identified as major actors of the EDPS paradigm. We conclude that the epigenetic dimension of a protein structure is an intrinsic weakness of AI-based protein structure prediction, especially AlphaFold, which warrants further development.

Keywords:  AI; alphafold; ganglioside; lipid rafts; membrane; molecular modeling; pathology; protein structure; therapy

DOI:  https://doi.org/10.3390/biom12101527
Front Bioinform. 2021 ;1 685844

Beyond Microsatellite Instability: Intrinsic Disorder as a Potential Link Between Protein Short Tandem Repeats and Cancer.

Max A Verbiest, Matteo Delucchi, Tugce Bilgin Sonay, Maria Anisimova.

  Short tandem repeats (STRs) are abundant in genomic sequences and are known for comparatively high mutation rates; STRs therefore are thought to be a potent source of genetic diversity. In protein-coding sequences STRs primarily encode disorder-promoting amino acids and are often located in intrinsically disordered regions (IDRs). STRs are frequently studied in the scope of microsatellite instability (MSI) in cancer, with little focus on the connection between protein STRs and IDRs. We believe, however, that this relationship should be explicitly included when ascertaining STR functionality in cancer. Here we explore this notion using all canonical human proteins from SwissProt, wherein we detected 3,699 STRs. Over 80% of these consisted completely of disorder promoting amino acids. 62.1% of amino acids in STR sequences were predicted to also be in an IDR, compared to 14.2% for non-repeat sequences. Over-representation analysis showed STR-containing proteins to be primarily located in the nucleus where they perform protein- and nucleotide-binding functions and regulate gene expression. They were also enriched in cancer-related signaling pathways. Furthermore, we found enrichments of STR-containing proteins among those correlated with patient survival for cancers derived from eight different anatomical sites. Intriguingly, several of these cancer types are not known to have a MSI-high (MSI-H) phenotype, suggesting that protein STRs play a role in cancer pathology in non MSI-H settings. Their intrinsic link with IDRs could therefore be an attractive topic of future research to further explore the role of STRs and IDRs in cancer. We speculate that our observations may be linked to the known dosage-sensitivity of disordered proteins, which could hint at a concentration-dependent gain-of-function mechanism in cancer for proteins containing STRs and IDRs.

Keywords:  cancer; computational biology; intrinsic disorder; microsatellite instability; microsatellites; protein bioinformatics; short tandem repeats

DOI:  https://doi.org/10.3389/fbinf.2021.685844
J Pers Med. 2022 Oct 06. pii: 1662. [Epub ahead of print]12(10):

Nanomedicines Meet Disordered Proteins: A Shift from Traditional Materials and Concepts to Innovative Polymers.

Bruno Rizzuti.

  Water-soluble nanomedicines have been widely studied for the targeted delivery of drugs for a very long time. As a notable example, biomaterials based on N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers have been under investigation for nearly half a century. In particular, anticancer drug carriers have been developed under the assumption that the leading mechanism with a therapeutic impact on solid tumors is the enhanced permeability and retention (EPR) effect, which dates back more than three decades. Nevertheless, these (and other) materials and concepts have encountered several barriers in their successful translation into clinical practice, and future nanomedicines need improvements in both passive and active targeting to their site of action. Notions borrowed from recent studies on intrinsically disordered proteins (IDPs) seem promising for enhancing the self-assembly, stimuli-responsiveness, and recognition properties of protein/peptide-based copolymers. Accordingly, IDP-based nanomedicines are ready to give new impetus to more traditional research in this field.

Keywords:  EPR; HPMA; copolymers; intrinsically disordered proteins; nanocarriers; nanomedicines

DOI:  https://doi.org/10.3390/jpm12101662
Biomolecules. 2022 Oct 19. pii: 1510. [Epub ahead of print]12(10):

The Effect of Positive Charge Distribution on the Cryoprotective Activity of Dehydrins.

Margaret A Smith, Steffen P Graether.

  Dehydrins are intrinsically disordered proteins expressed ubiquitously throughout the plant kingdom in response to desiccation. Dehydrins have been found to have a cryoprotective effect on lactate dehydrogenase (LDH) in vitro, which is in large part influenced by their hydrodynamic radius rather than the order of the amino acids within the sequence (alternatively, this may be a sequence specific effect). However, it seems that a different mechanism may underpin the cryoprotection that they confer to the cold-labile yeast frataxin homolog-1 (Yfh1). Circular dichroism spectroscopy (CD) was used to assess the degree of helicity of Yfh1 at 1 °C, both alone and in the presence of several dehydrin constructs. Three constructs were compared to the wild type: YSK2-K→R (lysine residues substituted with arginine), YSK2-Neutral (locally neutralized charge), and YSK2-SpaceK (evenly distributed positive charge). The results show that sequence rearrangements and minor substitutions have little impact on the ability of the dehydrin to preserve LDH activity. However, when the positive charge of the dehydrin is locally neutralized or evenly distributed, the dehydrin becomes less efficient at promoting structure in Yfh1 at low temperatures. This suggests that a stabilizing, charge-based interaction occurs between dehydrins and Yfh1. Dehydrins are intrinsically disordered proteins, expressed by certain organisms to improve desiccation tolerance. These proteins are thought to serve many cellular roles, such as the stabilization of membranes, DNA, and proteins. However, the molecular mechanisms underlying the function of dehydrins are not well understood. Here, we examine the importance of positive charges in dehydrin sequences by making substitutions and comparing their effects in the cryoprotection of two different proteins.

Keywords:  charge; circular dichroism; cryoprotection; dehydrins; intrinsically disordered proteins; lactate dehydrogenase; sequence order; yeast frataxin homolog 1

DOI:  https://doi.org/10.3390/biom12101510
Genes (Basel). 2022 Sep 22. pii: 1703. [Epub ahead of print]13(10):

Intrinsic Disorder in BAP1 and Its Association with Uveal Melanoma.

Mak B Djulbegovic, David J Taylor, Vladimir N Uversky, Anat Galor, Carol L Shields, Carol L Karp.

   BACKGROUND: Specific subvariants of uveal melanoma (UM) are associated with increased rates of metastasis compared to other subvariants. BRCA1 (BReast CAncer gene 1)-associated protein-1 (BAP1) is encoded by a gene that has been linked to aggressive behavior in UM.
METHODS: We evaluated BAP1 for the presence of intrinsically disordered protein regions (IDPRs) and its protein-protein interactions (PPI). We evaluated specific sequence-based features of the BAP1 protein using a set of bioinformatic databases, predictors, and algorithms.
RESULTS: We show that BAP1's structure contains extensive IDPRs as it is highly enriched in proline residues (the most disordered amino acid; p-value < 0.05), the average percent of predicted disordered residues (PPDR) was 57.34%, and contains 9 disorder-based binding sites (ie. molecular recognition features (MoRFs)). BAP1's intrinsic disorder allows it to engage in a complex PPI network with at least 49 partners (p-value < 1.0 × 10-16).
CONCLUSION: These findings show that BAP1 contains IDPRs and an intricate PPI network. Mutations in UM that are associated with the BAP1 gene may alter the function of the IDPRs embedded into its structure. These findings develop the understanding of UM and may provide a target for potential novel therapies to treat this aggressive neoplasm.

Keywords:  BAP1; intrinsically disordered protein (IDP); intrinsically disordered protein regions (IDPR); protein–protein interaction network; uveal melanoma

DOI:  https://doi.org/10.3390/genes13101703
Protein Sci. 2022 Nov;31(11): e4455

Intrinsic protein disorder uncouples affinity from binding specificity.

Tamas Lazar, Agnes Tantos, Peter Tompa, Eva Schad.

  Intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) of proteins often function by molecular recognition, in which they undergo induced folding. Based on prior generalizations, the idea prevails in the IDP field that due to the entropic penalty of induced folding, the major functional advantage associated with this binding mode is "uncoupling" specificity from binding strength. Nevertheless, both weaker binding and high specificity of IDPs/IDRs rest on limited experimental observations, making these assumptions more speculations than evidence-supported facts. The issue is also complicated by the rather vague concept of specificity that lacks an exact measure, such as the Kd for binding strength. We addressed these issues by creating and analyzing a comprehensive dataset of well-characterized ID/globular protein complexes, for which both the atomic structure of the complex and free energy (ΔG, Kd ) of interaction is known. Through this analysis, we provide evidence that the affinity distributions of IDP/globular and globular/globular complexes show different trends, whereas specificity does not connote to weaker binding strength of IDPs/IDRs. Furthermore, protein disorder extends the spectrum in the direction of very weak interactions, which may have important regulatory consequences and suggest that, in a biological sense, strict correlation of specificity and binding strength are uncoupled by structural disorder.

Keywords:  IDPs; binding strength; conservation; disordered protein complexes; specificity; structural disorder

DOI:  https://doi.org/10.1002/pro.4455
Front Bioinform. 2022 ;2 1019597

SETH predicts nuances of residue disorder from protein embeddings.

Dagmar Ilzhöfer, Michael Heinzinger, Burkhard Rost.

  Predictions for millions of protein three-dimensional structures are only a few clicks away since the release of AlphaFold2 results for UniProt. However, many proteins have so-called intrinsically disordered regions (IDRs) that do not adopt unique structures in isolation. These IDRs are associated with several diseases, including Alzheimer's Disease. We showed that three recent disorder measures of AlphaFold2 predictions (pLDDT, "experimentally resolved" prediction and "relative solvent accessibility") correlated to some extent with IDRs. However, expert methods predict IDRs more reliably by combining complex machine learning models with expert-crafted input features and evolutionary information from multiple sequence alignments (MSAs). MSAs are not always available, especially for IDRs, and are computationally expensive to generate, limiting the scalability of the associated tools. Here, we present the novel method SETH that predicts residue disorder from embeddings generated by the protein Language Model ProtT5, which explicitly only uses single sequences as input. Thereby, our method, relying on a relatively shallow convolutional neural network, outperformed much more complex solutions while being much faster, allowing to create predictions for the human proteome in about 1 hour on a consumer-grade PC with one NVIDIA GeForce RTX 3060. Trained on a continuous disorder scale (CheZOD scores), our method captured subtle variations in disorder, thereby providing important information beyond the binary classification of most methods. High performance paired with speed revealed that SETH's nuanced disorder predictions for entire proteomes capture aspects of the evolution of organisms. Additionally, SETH could also be used to filter out regions or proteins with probable low-quality AlphaFold2 3D structures to prioritize running the compute-intensive predictions for large data sets. SETH is freely publicly available at: https://github.com/Rostlab/SETH.

Keywords:  AlphaFold2; IDP; IDR; protein disorder; protein language model; protein structure prediction; residue disorder

DOI:  https://doi.org/10.3389/fbinf.2022.1019597
Cell Commun Signal. 2022 Oct 24. 20(1): 163

Does one plus one always equal two? Structural differences between nesfatin-1, -2, and nesfatin-1/2.

Rafał Lenda, Michał Padjasek, Artur Krężel, Andrzej Ożyhar, Dominika Bystranowska.

  Nesfatin-1 and -2 are produced from a reaction in which the N-terminus of human Nucleobindin-2 undergoes proteolytical processing. To date, Nucleobindin-2 and/or nesfatin-1 have only been shown to act as peptide hormones. On the other hand, the purpose of nesfatin-2 remains unknown. Since Nucleobindin-2/nesfatin-1 is thought impact the control of a wide range of physiological processes, including energy homeostasis, neurodegenerative processes and carcinogenesis, its ligands/interactions deserve special studies and attention. However, there are no reports about the molecular properties of the proteolytical products of human Nucleobindin-2 in the literature. Hence, this study aimed to analyze the effect of Zn(II) and Ca(II) on human nesfatin-1, -2, and -1/2 structures. Herein, we report that human nesfatin-1 is a member of the intrinsically disordered protein family, as indicated by circular dichroism and analytical ultracentrifugation experiments. In contrast, we found that the human nesfatin-2 and nesfatin-1/2 structures were globular with intrinsically disordered regions. Under Zn(II) treatment, we observed concentration-dependent structurization and compaction of intrinsically disordered nesfatin-1 and its propensity for oligomerization, as well as destabilization of both nesfatin-2 and nesfatin-1/2. Furthermore, dissociation constants for Zn(II) binding by nesfatin-1, nesfatin-2, and nesfatin-1/2 were also reported. Moreover, structurally distinct nesfatin-1 and -2 seem to be interdependent when linked together, as indicated by the observed molecular properties of nesfatin-1/2, which in turn are not a simple sum of the properties exhibited by the former peptides. Thus, herein, we shed new light on the molecular behavior of human nesfatins, which might help to elucidate the complex function of those peptides. Video abstract.

Keywords:  Hormone; IDP; Intrinsically disordered protein; Metalloprotein; Nesfatin-1; Nesfatin-2; Neuropeptide; Nucleobindin-2; Satiety molecule; Zinc

DOI:  https://doi.org/10.1186/s12964-022-00980-7
Biomolecules. 2022 Oct 15. pii: 1486. [Epub ahead of print]12(10):

Functional Tuning of Intrinsically Disordered Regions in Human Proteins by Composition Bias.

Kristina Kastano, Pablo Mier, Zsuzsanna Dosztányi, Vasilis J Promponas, Miguel A Andrade-Navarro.

  Intrinsically disordered regions (IDRs) in protein sequences are flexible, have low structural constraints and as a result have faster rates of evolution. This lack of evolutionary conservation greatly limits the use of sequence homology for the classification and functional assessment of IDRs, as opposed to globular domains. The study of IDRs requires other properties for their classification and functional prediction. While composition bias is not a necessary property of IDRs, compositionally biased regions (CBRs) have been noted as frequent part of IDRs. We hypothesized that to characterize IDRs, it could be helpful to study their overlap with particular types of CBRs. Here, we evaluate this overlap in the human proteome. A total of 2/3 of residues in IDRs overlap CBRs. Considering CBRs enriched in one type of amino acid, we can distinguish CBRs that tend to be fully included within long IDRs (R, H, N, D, P, G), from those that partially overlap shorter IDRs (S, E, K, T), and others that tend to overlap IDR terminals (Q, A). CBRs overlap more often IDRs in nuclear proteins and in proteins involved in liquid-liquid phase separation (LLPS). Study of protein interaction networks reveals the enrichment of CBRs in IDRs by tandem repetition of short linear motifs (rich in S or P), and the existence of E-rich polar regions that could support specific protein interactions with non-specific interactions. Our results open ways to pin down the function of IDRs from their partial compositional biases.

Keywords:  compositionally biased regions; intrinsically disordered regions; liquid–liquid phase separation; low complexity regions

DOI:  https://doi.org/10.3390/biom12101486
Proteins. 2022 Oct 28.

A sequence-based foldability score combined with AlphaFold2 predictions to disentangle the protein order/disorder continuum.

Apolline Bruley, Tristan Bitard-Feildel, Isabelle Callebaut, Elodie Duprat.

  Order and disorder govern protein functions, but there is a great diversity in disorder, from regions that are - and stay - fully disordered to conditional order. This diversity is still difficult to decipher even though it is encoded in the amino acid sequences. Here, we developed an analytic Python package, named pyHCA, to estimate the foldability of a protein segment from the only information of its amino acid sequence and based on a measure of its density in regular secondary structures associated with hydrophobic clusters, as defined by the Hydrophobic Cluster Analysis (HCA) approach. The tool was designed by optimizing the separation between foldable segments from databases of disorder (DisProt) and order (SCOPe (soluble domains) and OPM (transmembrane domains)). It allows to specify the ratio between order, embodied by regular secondary structures (either participating in the hydrophobic core of well-folded 3D structures or conditionally formed in intrinsically disordered regions) and disorder. We illustrated the relevance of pyHCA with several examples and applied it to the sequences of the proteomes of 21 species ranging from prokaryotes and archaea to unicellular and multicellular eukaryotes, for which structure models are provided in the AlphaFold2 databases. Cases of low-confidence scores related to disorder were distinguished from those of sequences that we identified as foldable but are still excluded from accurate modeling by AlphaFold2 due to a lack of sequence homologs or to compositional biases. Overall, our approach is complementary to AlphaFold2, providing guides to map structural innovations through evolutionary processes, at proteome and gene scales. This article is protected by copyright. All rights reserved.

Keywords:  AlphaFold protein structure database; Hydrophobic Cluster Analysis; IDPs/IDRs; Protein foldable segments; soluble and transmembrane domains

DOI:  https://doi.org/10.1002/prot.26441
Biomolecules. 2022 Oct 08. pii: 1441. [Epub ahead of print]12(10):

Reorganization of Cell Compartmentalization Induced by Stress.

Anna S Fefilova, Iuliia A Antifeeva, Anastasia A Gavrilova, Konstantin K Turoverov, Irina M Kuznetsova, Alexander V Fonin.

  The discovery of intrinsically disordered proteins (IDPs) that do not have an ordered structure and nevertheless perform essential functions has opened a new era in the understanding of cellular compartmentalization. It threw the bridge from the mostly mechanistic model of the organization of the living matter to the idea of highly dynamic and functional "soft matter". This paradigm is based on the notion of the major role of liquid-liquid phase separation (LLPS) of biopolymers in the spatial-temporal organization of intracellular space. The LLPS leads to the formation of self-assembled membrane-less organelles (MLOs). MLOs are multicomponent and multifunctional biological condensates, highly dynamic in structure and composition, that allow them to fine-tune the regulation of various intracellular processes. IDPs play a central role in the assembly and functioning of MLOs. The LLPS importance for the regulation of chemical reactions inside the cell is clearly illustrated by the reorganization of the intracellular space during stress response. As a reaction to various types of stresses, stress-induced MLOs appear in the cell, enabling the preservation of the genetic and protein material during unfavourable conditions. In addition, stress causes structural, functional, and compositional changes in the MLOs permanently present inside the cells. In this review, we describe the assembly of stress-induced MLOs and the stress-induced modification of existing MLOs in eukaryotes, yeasts, and prokaryotes in response to various stress factors.

Keywords:  intrinsically disordered proteins; liquid-liquid phase separation; membrane-less organelles; stress

DOI:  https://doi.org/10.3390/biom12101441