A Family of Small Intrinsically Disordered Proteins Involved in Flagellum-Dependent Motility inSalmonella enterica

ABSTRACTBy screening a collection ofSalmonellamutants deleted for genes encoding small proteins of ≤60 amino acids, we identified three paralogous small genes (ymdF,STM14_1829, andyciG) required for wild-type flagellum-dependent swimming and swarming motility. TheymdF,STM14_1829, andyciGgenes encode small proteins of 55, 60, and 60 amino acid residues, respectively. A bioinformatics analysis predicted that these small proteins are intrinsically disordered proteins, and circular dichroism analysis of purified recombinant proteins confirmed that all three proteins are unstructured in solution. A mutant deleted for STM14_1829 showed the most severe motility defect, indicating that among the three paralogs, STM14_1829 is a key protein required for wild-type motility. We determined that relative to the wild type, the expression of the flagellin protein FliC is lower in the ΔSTM14_1829mutant due to the downregulation of theflhDCoperon encoding the FlhDC master regulator. By comparing the gene expression profiles between the wild-type and ΔSTM14_1829strains via RNA sequencing, we found that the gene encoding the response regulator PhoP is upregulated in the ΔSTM14_1829mutant, suggesting the indirect repression of theflhDCoperon by the activated PhoP. Homologs of STM14_1829 are conserved in a wide range of bacteria, includingEscherichia coliandPseudomonas aeruginosa. We showed that the inactivation of STM14_1829 homologs inE. coliandP. aeruginosaalso alters motility, suggesting that this family of small intrinsically disordered proteins may play a role in the cellular pathway(s) that affects motility.IMPORTANCEThis study reports the identification of a novel family of small intrinsically disordered proteins that are conserved in a wide range of flagellated and nonflagellated bacteria. Although this study identifies the role of these small proteins in the scope of flagellum-dependent motility inSalmonella, they likely play larger roles in a more conserved cellular pathway(s) that indirectly affects flagellum expression in the case of motile bacteria. Small intrinsically disordered proteins have not been well characterized in prokaryotes, and the results of our study provide a basis for their detailed functional characterization.

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Intrinsically disordered caldesmon binds calmodulin via the “buttons on a string” mechanism

PeerJ ◽

10.7717/peerj.1265 ◽

2015 ◽

Vol 3 ◽

pp. e1265 ◽

Cited By ~ 6

Author(s):

Sergei E. Permyakov ◽

Eugene A. Permyakov ◽

Vladimir N. Uversky

Keyword(s):

Local Structure ◽

Intrinsically Disordered Proteins ◽

String Model ◽

Electrostatic Attraction ◽

Disordered Proteins ◽

Wild Type ◽

Intrinsically Disordered ◽

Tryptophan Residues ◽

Specific Packing ◽

Oppositely Charged

We show here that chicken gizzard caldesmon (CaD) and its C-terminal domain (residues 636–771, CaD136) are intrinsically disordered proteins. The computational and experimental analyses of the wild type CaD136and series of its single tryptophan mutants (W674A, W707A, and W737A) and a double tryptophan mutant (W674A/W707A) suggested that although the interaction of CaD136with calmodulin (CaM) can be driven by the non-specific electrostatic attraction between these oppositely charged molecules, the specificity of CaD136-CaM binding is likely to be determined by the specific packing of important CaD136tryptophan residues at the CaD136-CaM interface. It is suggested that this interaction can be described as the “buttons on a charged string” model, where the electrostatic attraction between the intrinsically disordered CaD136and the CaM is solidified in a “snapping buttons” manner by specific packing of the CaD136“pliable buttons” (which are the short segments of fluctuating local structure condensed around the tryptophan residues) at the CaD136-CaM interface. Our data also show that all three “buttons” are important for binding, since mutation of any of the tryptophans affects CaD136-CaM binding and since CaD136remains CaM-buttoned even when two of the three tryptophans are mutated to alanines.

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Supramolecular Fuzziness of Intracellular Liquid Droplets: Liquid–Liquid Phase Transitions, Membrane-Less Organelles, and Intrinsic Disorder

Molecules ◽

10.3390/molecules24183265 ◽

2019 ◽

Vol 24 (18) ◽

pp. 3265 ◽

Cited By ~ 12

Author(s):

Vladimir N. Uversky

Keyword(s):

Phase Transitions ◽

Liquid Phase ◽

Intrinsically Disordered Proteins ◽

Intrinsically Disordered Protein ◽

Disordered Proteins ◽

Bacterial Cells ◽

Liquid Droplets ◽

Intrinsically Disordered ◽

Wide Range ◽

Characteristic Features

Cells are inhomogeneously crowded, possessing a wide range of intracellular liquid droplets abundantly present in the cytoplasm of eukaryotic and bacterial cells, in the mitochondrial matrix and nucleoplasm of eukaryotes, and in the chloroplast’s stroma of plant cells. These proteinaceous membrane-less organelles (PMLOs) not only represent a natural method of intracellular compartmentalization, which is crucial for successful execution of various biological functions, but also serve as important means for the processing of local information and rapid response to the fluctuations in environmental conditions. Since PMLOs, being complex macromolecular assemblages, possess many characteristic features of liquids, they represent highly dynamic (or fuzzy) protein–protein and/or protein–nucleic acid complexes. The biogenesis of PMLOs is controlled by specific intrinsically disordered proteins (IDPs) and hybrid proteins with ordered domains and intrinsically disordered protein regions (IDPRs), which, due to their highly dynamic structures and ability to facilitate multivalent interactions, serve as indispensable drivers of the biological liquid–liquid phase transitions (LLPTs) giving rise to PMLOs. In this article, the importance of the disorder-based supramolecular fuzziness for LLPTs and PMLO biogenesis is discussed.

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Chasing coevolutionary signals in intrinsically disordered proteins complexes

Scientific Reports ◽

10.1038/s41598-020-74791-6 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Javier A. Iserte ◽

Tamas Lazar ◽

Silvio C. E. Tosatto ◽

Peter Tompa ◽

Cristina Marino-Buslje

Keyword(s):

Protein Interactions ◽

Intrinsically Disordered Proteins ◽

Disordered Proteins ◽

Protein Protein Interactions ◽

Contact Prediction ◽

Intrinsically Disordered ◽

Regulatory Processes ◽

Wide Range ◽

Predict Protein Structure ◽

Biological Interpretation

Abstract Intrinsically disordered proteins/regions (IDPs/IDRs) are crucial components of the cell, they are highly abundant and participate ubiquitously in a wide range of biological functions, such as regulatory processes and cell signaling. Many of their important functions rely on protein interactions, by which they trigger or modulate different pathways. Sequence covariation, a powerful tool for protein contact prediction, has been applied successfully to predict protein structure and to identify protein–protein interactions mostly of globular proteins. IDPs/IDRs also mediate a plethora of protein–protein interactions, highlighting the importance of addressing sequence covariation-based inter-protein contact prediction of this class of proteins. Despite their importance, a systematic approach to analyze the covariation phenomena of intrinsically disordered proteins and their complexes is still missing. Here we carry out a comprehensive critical assessment of coevolution-based contact prediction in IDP/IDR complexes and detail the challenges and possible limitations that emerge from their analysis. We found that the coevolutionary signal is faint in most of the complexes of disordered proteins but positively correlates with the interface size and binding affinity between partners. In addition, we discuss the state-of-art methodology by biological interpretation of the results, formulate evaluation guidelines and suggest future directions of development to the field.

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Ion Mobility Mass Spectrometry Uncovers the Impact of the Patterning of Oppositely Charged Residues on the Conformational Distributions of Intrinsically Disordered Proteins

10.26434/chemrxiv.7312277.v1 ◽

2018 ◽

Cited By ~ 2

Author(s):

Rebecca Beveridge ◽

Lukasz Migas ◽

Rahul Das ◽

Rohit Pappu ◽

Richard Kriwacki ◽

...

Keyword(s):

Mass Spectrometry ◽

Ion Mobility ◽

Intrinsically Disordered Proteins ◽

Disordered Proteins ◽

Ion Mobility Mass Spectrometry ◽

Conformational Heterogeneity ◽

Wild Type ◽

Intrinsically Disordered ◽

Charged Residues ◽

Oppositely Charged

The global dimensions and amplitudes of conformational fluctuations of intrinsically disordered proteins are governed, in part, by the linear segregation versus clustering of oppositely charged residues within the primary sequence. Ion Mobility-Mass Spectrometry (IM-MS) affords unique advantages for probing the conformational consequences of the linear patterning of oppositely charged residues because it measures and separates proteins electrosprayed from solution on the basis of charge and shape. Here, we use IM-MS to measure the conformational consequences of charge patterning on the C-terminal intrinsically disordered region (p27 IDR) of the cell cycle inhibitory protein p27<sup>Kip1</sup>. We report the range of charge states and accompanying collisional cross section distributions for wild-type p27 IDR and two variants with identical amino acid compositions, k14 and k56, distinguished by the extent of linear mixing versus segregation of oppositely charged residues. Wild-type p27 IDR (k31) and k14 where the oppositely charged residues are more evenly distributed, exhibit a broad distribution of charge states. This is concordant with high degrees of conformational heterogeneity in solution. By contrast, k56 with linear segregation of oppositely charged residues, leads to limited conformational heterogeneity and a narrow distribution of charged states. Molecular dynamics simulations demonstrate that the interplay between chain solvation and intra-chain interactions (self-solvation) leads to conformational distributions that are modulated by salt concentration, with the wild-type sequence showing the most sensitivity to changes in salt concentration. These results suggest that the charge patterning within the wild-type p27 IDR may be optimized to sample both highly solvated and self-solvated conformational states.

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Conserved Eukaryotic Kinase CK2 Chaperone Intrinsically Disordered Protein Interactions

Applied and Environmental Microbiology ◽

10.1128/aem.02191-19 ◽

2019 ◽

Vol 86 (2) ◽

Author(s):

Lianhu Zhang ◽

Dongmei Zhang ◽

Dan Liu ◽

Yuan Li ◽

Hongchen Li ◽

...

Keyword(s):

Fusarium Graminearum ◽

Intrinsically Disordered Proteins ◽

Large Range ◽

Disordered Proteins ◽

Interacting Proteins ◽

General Role ◽

Content Type ◽

Intrinsically Disordered ◽

Dna And Rna ◽

Aggregation And Disaggregation

ABSTRACT CK2, a serine/threonine (Ser/Thr) kinase present in eukaryotic cells, is known to have a vast number of substrates. We have recently shown that it localizes to nuclei and at pores between hyphal compartments in Magnaporthe oryzae. We performed a pulldown proteomics analysis of M. oryzae CK2 catalytic subunit MoCKa to detect interacting proteins. The MoCKa pulldown was enriched for septum and nucleolus proteins and intrinsically disordered proteins (IDPs) containing a CK2 phosphorylation motif that is proposed to destabilize and unfold α-helices. This points to a function for CK2 phosphorylation and corresponding phosphatase dephosphorylation in the formation of functional protein-protein aggregates and protein-RNA/DNA binding. To test this as widely as possible, we used secondary data downloaded from databases from a large range of M. oryzae experiments, as well as data for a relatively closely related plant-pathogenic fungus, Fusarium graminearum. We found that CKa expression was strongly positively correlated with Ser/Thr phosphatases, as well as with disaggregases (HSP104, YDJ1, and SSA1) and an autophagy-indicating protein (ATG8). The latter points to increased protein aggregate formation at high levels of CKa expression. Our results suggest a general role for CK2 in chaperoning aggregation and disaggregation of IDPs and their binding to proteins, DNA, and RNA. IMPORTANCE CK2 is a eukaryotic conserved kinase enzyme complex that phosphorylates proteins. CK2 is known to phosphorylate a large number of proteins and is constitutively active, and thus a “normal” role for a kinase in a signaling cascade might not be the case for CK2. Previous results on localization and indications from the literature point to a function for CK2 phosphorylation in shaping and folding of proteins, especially intrinsically disordered proteins, which constitute about 30% of eukaryotic proteins. We used pulldown of interacting proteins and data downloaded from a large range of transcriptomic experiments in M. oryzae and complemented these with data downloaded from a large range of transcriptomic experiments in Fusarium graminearum. We found support for a general role for CK2 in aggregation and disaggregation of IDPs and their binding to proteins, DNA, and RNA—interactions that could explain the importance of CK2 in eukaryotic cell function and disease.

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On the roles of intrinsically disordered proteins and regions in cell communication and signaling

Cell Communication and Signaling ◽

10.1186/s12964-021-00774-3 ◽

2021 ◽

Vol 19 (1) ◽

Cited By ~ 1

Author(s):

Sarah E. Bondos ◽

A. Keith Dunker ◽

Vladimir N. Uversky

Keyword(s):

Intrinsically Disordered Proteins ◽

Cell Communication ◽

Disordered Proteins ◽

Sequence Structure ◽

Intrinsically Disordered ◽

Wide Range ◽

Micro Organisms ◽

Signaling Domains ◽

Structured Proteins ◽

Water Bears

AbstractFor proteins, the sequence → structure → function paradigm applies primarily to enzymes, transmembrane proteins, and signaling domains. This paradigm is not universal, but rather, in addition to structured proteins, intrinsically disordered proteins and regions (IDPs and IDRs) also carry out crucial biological functions. For these proteins, the sequence → IDP/IDR ensemble → function paradigm applies primarily to signaling and regulatory proteins and regions. Often, in order to carry out function, IDPs or IDRs cooperatively interact, either intra- or inter-molecularly, with structured proteins or other IDPs or intermolecularly with nucleic acids. In this IDP/IDR thematic collection published in Cell Communication and Signaling, thirteen articles are presented that describe IDP/IDR signaling molecules from a variety of organisms from humans to fruit flies and tardigrades (“water bears”) and that describe how these proteins and regions contribute to the function and regulation of cell signaling. Collectively, these papers exhibit the diverse roles of disorder in responding to a wide range of signals as to orchestrate an array of organismal processes. They also show that disorder contributes to signaling in a broad spectrum of species, ranging from micro-organisms to plants and animals.

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The dynein light chain 8 (LC8) binds predominantly “in-register” to a multivalent intrinsically disordered partner

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.011653 ◽

2020 ◽

Vol 295 (15) ◽

pp. 4912-4922 ◽

Cited By ~ 2

Author(s):

Patrick N. Reardon ◽

Kayla A. Jara ◽

Amber D. Rolland ◽

Delaney A. Smith ◽

Hanh T. M. Hoang ◽

...

Keyword(s):

Light Chain ◽

Intrinsically Disordered Proteins ◽

Analytical Ultracentrifugation ◽

Complex Dynamics ◽

Regulatory Protein ◽

Disordered Proteins ◽

Dynein Light Chain ◽

Linker Length ◽

Intrinsically Disordered ◽

Wide Range

Dynein light chain 8 (LC8) interacts with intrinsically disordered proteins (IDPs) and influences a wide range of biological processes. It is becoming apparent that among the numerous IDPs that interact with LC8, many contain multiple LC8-binding sites. Although it is established that LC8 forms parallel IDP duplexes with some partners, such as nucleoporin Nup159 and dynein intermediate chain, the molecular details of these interactions and LC8's interactions with other diverse partners remain largely uncharacterized. LC8 dimers could bind in either a paired “in-register” or a heterogeneous off-register manner to any of the available sites on a multivalent partner. Here, using NMR chemical shift perturbation, analytical ultracentrifugation, and native electrospray ionization MS, we show that LC8 forms a predominantly in-register complex when bound to an IDP domain of the multivalent regulatory protein ASCIZ. Using saturation transfer difference NMR, we demonstrate that at substoichiometric LC8 concentrations, the IDP domain preferentially binds to one of the three LC8 recognition motifs. Further, the differential dynamic behavior for the three sites and the size of the fully bound complex confirmed an in-register complex. Dynamics measurements also revealed that coupling between sites depends on the linker length separating these sites. These results identify linker length and motif specificity as drivers of in-register binding in the multivalent LC8–IDP complex assembly and the degree of compositional and conformational heterogeneity as a promising emerging mechanism for tuning of binding and regulation.

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The Prediction of Intrinsically Disordered Proteins Based on Feature Selection

Algorithms ◽

10.3390/a12020046 ◽

2019 ◽

Vol 12 (2) ◽

pp. 46 ◽

Cited By ~ 2

Author(s):

Hao He ◽

Jiaxiang Zhao ◽

Guiling Sun

Keyword(s):

Intrinsically Disordered Proteins ◽

Back Propagation ◽

Disordered Proteins ◽

Evolutionary Information ◽

Multi Layer Perceptron ◽

Biological Functions ◽

Training Process ◽

Intrinsically Disordered ◽

Wide Range ◽

Simulation Results

Intrinsically disordered proteins perform a variety of important biological functions, which makes their accurate prediction useful for a wide range of applications. We develop a scheme for predicting intrinsically disordered proteins by employing 35 features including eight structural properties, seven physicochemical properties and 20 pieces of evolutionary information. In particular, the scheme includes a preprocessing procedure which greatly reduces the input features. Using two different windows, the preprocessed data containing not only the properties of the surroundings of the target residue but also the properties related to the specific target residue are fed into a multi-layer perceptron neural network as its inputs. The Adam algorithm for the back propagation together with the dropout algorithm to avoid overfitting are introduced during the training process. The training as well as testing our procedure is performed on the dataset DIS803 from a DisProt database. The simulation results show that the performance of our scheme is competitive in comparison with ESpritz and IsUnstruct.

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Protein disorder-to-order transition enhances the nucleosome-binding affinity of H1

Nucleic Acids Research ◽

10.1093/nar/gkaa285 ◽

2020 ◽

Vol 48 (10) ◽

pp. 5318-5331 ◽

Cited By ~ 4

Author(s):

Akshay Sridhar ◽

Modesto Orozco ◽

Rosana Collepardo-Guevara

Keyword(s):

Binding Affinity ◽

Binding Proteins ◽

Intrinsically Disordered Proteins ◽

Order Transition ◽

Disordered Proteins ◽

Chromatin Binding ◽

Intrinsically Disordered ◽

Terminal Domain ◽

Wide Range ◽

Nucleosome Binding

Abstract Intrinsically disordered proteins are crucial elements of chromatin heterogenous organization. While disorder in the histone tails enables a large variation of inter-nucleosome arrangements, disorder within the chromatin-binding proteins facilitates promiscuous binding to a wide range of different molecular targets, consistent with structural heterogeneity. Among the partially disordered chromatin-binding proteins, the H1 linker histone influences a myriad of chromatin characteristics including compaction, nucleosome spacing, transcription regulation, and the recruitment of other chromatin regulating proteins. Although it is now established that the long C-terminal domain (CTD) of H1 remains disordered upon nucleosome binding and that such disorder favours chromatin fluidity, the structural behaviour and thereby the role/function of the N-terminal domain (NTD) within chromatin is yet unresolved. On the basis of microsecond-long parallel-tempering metadynamics and temperature-replica exchange atomistic molecular dynamics simulations of different H1 NTD subtypes, we demonstrate that the NTD is completely unstructured in solution but undergoes an important disorder-to-order transition upon nucleosome binding: it forms a helix that enhances its DNA binding ability. Further, we show that the helical propensity of the H1 NTD is subtype-dependent and correlates with the experimentally observed binding affinity of H1 subtypes, suggesting an important functional implication of this disorder-to-order transition.

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Does intrinsically disordered caldesmon bind calmodulin via the “buttons on a string” mechanism?

10.7287/peerj.preprints.1261v1 ◽

2015 ◽

Author(s):

Sergei E Permyakov ◽

Eugene A Permyakov ◽

Vladimir N Uversky

Keyword(s):

Local Structure ◽

Intrinsically Disordered Proteins ◽

String Model ◽

Electrostatic Attraction ◽

Disordered Proteins ◽

Wild Type ◽

Intrinsically Disordered ◽

Tryptophan Residues ◽

Specific Packing ◽

Oppositely Charged

We show here that chicken gizzard caldesmon (CaD) and its C-terminal domain (residues 636-771, CaD136) are intrinsically disordered proteins. The computational and experimental analyses of the wild type CaD136 and series of its single tryptophan mutants (W674A, W707A, and W737A) and a double tryptophan mutant (W674A/W707A) suggested that although the interaction of CaD136 with calmodulin (CaM) can be driven by the non-specific electrostatic attraction between these oppositely charged molecules, the specificity of CaD136-CaM binding is likely to be determined by the specific packing of important CaD136 tryptophan residues at the CaD136-CaM interface. It is suggested that this interaction can be described as the “buttons on a charged string” model, where the electrostatic attraction between the intrinsically disordered CaD136 and the CaM is solidified in a “snapping buttons” manner by specific packing of the CaD136 “pliable buttons” (which are the short segments of fluctuating local structure condensed around the tryptophan residues) at the CaD136-CaM interface. Our data also show that all three “buttons” are important for binding, since mutation of any of the tryptophans affects CaD136-CaM binding and since CaD136 remains CaM-buttoned even when two of the three tryptophans are mutated to alanines.

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