Christopher J. Oldfield

Intrinsically Disordered Proteins in Human Diseases: Introducing the D2 Concept

Intrinsically disordered proteins (IDPs) lack stable tertiary and/or secondary structures under physiological conditions in vitro. They are highly abundant in nature and their functional repertoire complements the functions of ordered proteins. IDPs are involved in regulation, signaling, and control, where binding to multiple partners and high-specificity/low-affinity interactions play a crucial role. Functions of IDPs are tuned via alternative splicing and posttranslational modifications. Intrinsic disorder is a unique structural feature that enables IDPs to participate in both one-to-many and many-to-one signaling. Numerous IDPs are associated with human diseases, including cancer, cardiovascular disease, amyloidoses, neurodegenerative diseases, and diabetes. Overall, intriguing interconnections among intrinsic disorder, cell signaling, and human diseases suggest that protein conformational diseases may result not only from protein misfolding, but also from misidentification, missignaling, and unnatural or nonnative folding. IDPs, such as alpha-synuclein, tau protein, p53, and BRCA1, are attractive targets for drugs modulating protein-protein interactions. From these and other examples, novel strategies for drug discovery based on IDPs have been developed. To summarize work in this area, we are introducing the D2 (disorder in disorders) concept.

Classification of Intrinsically Disordered Regions and Proteins.

1.1. Uncharacterized Protein Segments Are a Source of Functional Novelty Over the past decade, we have observed a massive increase in the amount of information describing protein sequences from a variety of organisms.1,2 While this may reflect the diversity in sequence space, and possibly also in function space,3 a large proportion of the sequences lacks any useful function annotation.4,5 Often these sequences are annotated as putative or hypothetical proteins, and for the majority their functions still remain unknown.6,7 Suggestions about potential protein function, primarily molecular function, often come from computational analysis of their sequences. For instance, homology detection allows for the transfer of information from well-characterized protein segments to those with similar sequences that lack annotation of molecular function.8−10 Other aspects of function, such as the biological processes proteins participate in, may come from genetic- and disease-association studies, expression and interaction network data, and comparative genomics approaches that investigate genomic context.11−17 Characterization of unannotated and uncharacterized protein segments is expected to lead to the discovery of novel functions as well as provide important insights into existing biological processes. In addition, it is likely to shed new light on molecular mechanisms of diseases that are not yet fully understood. Thus, uncharacterized protein segments are likely to be a large source of functional novelty relevant for discovering new biology.

Intrinsic disorder is a common feature of hub proteins from four eukaryotic interactomes.

Recent proteome-wide screening approaches have provided a wealth of information about interacting proteins in various organisms. To test for a potential association between protein connectivity and the amount of predicted structural disorder, the disorder propensities of proteins with various numbers of interacting partners from four eukaryotic organisms (Caenorhabditis elegans, Saccharomyces cerevisiae, Drosophila melanogaster, and Homo sapiens) were investigated. The results of PONDR VL-XT disorder analysis show that for all four studied organisms, hub proteins, defined here as those that interact with ≥10 partners, are significantly more disordered than end proteins, defined here as those that interact with just one partner. The proportion of predicted disordered residues, the average disorder score, and the number of predicted disordered regions of various lengths were higher overall in hubs than in ends. A binary classification of hubs and ends into ordered and disordered subclasses using the consensus prediction method showed a significant enrichment of wholly disordered proteins and a significant depletion of wholly ordered proteins in hubs relative to ends in worm, fly, and human. The functional annotation of yeast hubs and ends using GO categories and the correlation of these annotations with disorder predictions demonstrate that proteins with regulation, transcription, and development annotations are enriched in disorder, whereas proteins with catalytic activity, transport, and membrane localization annotations are depleted in disorder. The results of this study demonstrate that intrinsic structural disorder is a distinctive and common characteristic of eukaryotic hub proteins, and that disorder may serve as a determinant of protein interactivity.

Flexible nets: disorder and induced fit in the associations of p53 and 14-3-3 with their partners

BackgroundProteins are involved in many interactions with other proteins leading to networks that regulate and control a wide variety of physiological processes. Some of these proteins, called hub proteins or hubs, bind to many different protein partners. Protein intrinsic disorder, via diversity arising from structural plasticity or flexibility, provide a means for hubs to associate with many partners (Dunker AK, Cortese MS, Romero P, Iakoucheva LM, Uversky VN: Flexible Nets: The roles of intrinsic disorder in protein interaction networks. FEBS J 2005, 272:5129-5148).ResultsHere we present a detailed examination of two divergent examples: 1) p53, which uses different disordered regions to bind to different partners and which also has several individual disordered regions that each bind to multiple partners, and 2) 14-3-3, which is a structured protein that associates with many different intrinsically disordered partners. For both examples, three-dimensional structures of multiple complexes reveal that the flexibility and plasticity of intrinsically disordered protein regions as well as induced-fit changes in the structured regions are both important for binding diversity.ConclusionsThese data support the conjecture that hub proteins often utilize intrinsic disorder to bind to multiple partners and provide detailed information about induced fit in structured regions.

Evolutionary rate heterogeneity in proteins with long disordered regions

The dominant view in protein science is that a three-dimensional (3-D) structure is a prerequisite for protein function. In contrast to this dominant view, there are many counterexample proteins that fail to fold into a 3-D structure, or that have local regions that fail to fold, and yet carry out function. Protein without fixed 3-D structure is called intrinsically disordered. Motivated by anecdotal accounts of higher rates of sequence evolution in disordered protein than in ordered protein we are exploring the molecular evolution of disordered proteins. To test whether disordered protein evolves more rapidly than ordered protein, pairwise genetic distances were compared between the ordered and the disordered regions of 26 protein families having at least one member with a structurally characterized region of disorder of 30 or more consecutive residues. For five families, there were no significant differences in pairwise genetic distances between ordered and disordered sequences. The disordered region evolved significantly more rapidly than the ordered region for 19 of the 26 families. The functions of these disordered regions are diverse, including binding sites for protein, DNA, or RNA and also including flexible linkers. The functions of some of these regions are unknown. The disordered regions evolved significantly more slowly than the ordered regions for the two remaining families. The functions of these more slowly evolving disordered regions include sites for DNA binding. More work is needed to understand the underlying causes of the variability in the evolutionary rates of intrinsically ordered and disordered protein.

Intrinsically Disordered Proteins and Intrinsically Disordered Protein Regions

Intrinsically disordered proteins (IDPs) and IDP regions fail to form a stable structure, yet they exhibit biological activities. Their mobile flexibility and structural instability are encoded by their amino acid sequences. They recognize proteins, nucleic acids, and other types of partners; they accelerate interactions and chemical reactions between bound partners; and they help accommodate posttranslational modifications, alternative splicing, protein fusions, and insertions or deletions. Overall, IDP-associated biological activities complement those of structured proteins. Recently, there has been an explosion of studies on IDP regions and their functions, yet the discovery and investigation of these proteins have a long, mostly ignored history. Along with recent discoveries, we present several early examples and the mechanisms by which IDPs contribute to function, which we hope will encourage comprehensive discussion of IDPs and IDP regions in biochemistry textbooks. Finally, we propose future directions for IDP research.

Close encounters of the third kind: disordered domains and the interactions of proteins

Protein–protein interactions are thought to be mediated by domains, which are autonomous folding units of proteins. Recently, a second type of interaction has been suggested, mediated by short segments termed linear motifs, which are related to recognition elements of intrinsically disordered regions. Here, we propose a third kind of protein–protein recognition mechanism, mediated by disordered regions longer than 20–30 residues. Bioinformatics predictions and well‐characterized examples, such as the kinase‐inhibitory domain of Cdk inhibitors and the Wiskott–Aldrich syndrome protein (WASP)‐homology domain 2 of actin‐binding proteins, show that these disordered regions conform to the definition of domains rather than motifs, i.e., they represent functional, evolutionary, and structural units. Their functions are distinct from those of short motifs and ordered domains, and establish a third kind of interaction principle. With these points, we argue that these long disordered regions should be recognized as a distinct class of biologically functional protein domains.

Unfoldomics of human diseases: linking protein intrinsic disorder with diseases

BackgroundIntrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs) lack stable tertiary and/or secondary structure yet fulfills key biological functions. The recent recognition of IDPs and IDRs is leading to an entire field aimed at their systematic structural characterization and at determination of their mechanisms of action. Bioinformatics studies showed that IDPs and IDRs are highly abundant in different proteomes and carry out mostly regulatory functions related to molecular recognition and signal transduction. These activities complement the functions of structured proteins. IDPs and IDRs were shown to participate in both one-to-many and many-to-one signaling. Alternative splicing and posttranslational modifications are frequently used to tune the IDP functionality. Several individual IDPs were shown to be associated with human diseases, such as cancer, cardiovascular disease, amyloidoses, diabetes, neurodegenerative diseases, and others. This raises questions regarding the involvement of IDPs and IDRs in various diseases.ResultsIDPs and IDRs were shown to be highly abundant in proteins associated with various human maladies. As the number of IDPs related to various diseases was found to be very large, the concepts of the disease-related unfoldome and unfoldomics were introduced. Novel bioinformatics tools were proposed to populate and characterize the disease-associated unfoldome. Structural characterization of the members of the disease-related unfoldome requires specialized experimental approaches. IDPs possess a number of unique structural and functional features that determine their broad involvement into the pathogenesis of various diseases.ConclusionProteins associated with various human diseases are enriched in intrinsic disorder. These disease-associated IDPs and IDRs are real, abundant, diversified, vital, and dynamic. These proteins and regions comprise the disease-related unfoldome, which covers a significant part of the human proteome. Profound association between intrinsic disorder and various human diseases is determined by a set of unique structural and functional characteristics of IDPs and IDRs. Unfoldomics of human diseases utilizes unrivaled bioinformatics and experimental techniques, paves the road for better understanding of human diseases, their pathogenesis and molecular mechanisms, and helps develop new strategies for the analysis of disease-related proteins.

MoRFpred, a computational tool for sequence-based prediction and characterization of short disorder-to-order transitioning binding regions in proteins

Motivation: Molecular recognition features (MoRFs) are short binding regions located within longer intrinsically disordered regions that bind to protein partners via disorder-to-order transitions. MoRFs are implicated in important processes including signaling and regulation. However, only a limited number of experimentally validated MoRFs is known, which motivates development of computational methods that predict MoRFs from protein chains. Results: We introduce a new MoRF predictor, MoRFpred, which identifies all MoRF types (α, β, coil and complex). We develop a comprehensive dataset of annotated MoRFs to build and empirically compare our method. MoRFpred utilizes a novel design in which annotations generated by sequence alignment are fused with predictions generated by a Support Vector Machine (SVM), which uses a custom designed set of sequence-derived features. The features provide information about evolutionary profiles, selected physiochemical properties of amino acids, and predicted disorder, solvent accessibility and B-factors. Empirical evaluation on several datasets shows that MoRFpred outperforms related methods: α-MoRF-Pred that predicts α-MoRFs and ANCHOR which finds disordered regions that become ordered when bound to a globular partner. We show that our predicted (new) MoRF regions have non-random sequence similarity with native MoRFs. We use this observation along with the fact that predictions with higher probability are more accurate to identify putative MoRF regions. We also identify a few sequence-derived hallmarks of MoRFs. They are characterized by dips in the disorder predictions and higher hydrophobicity and stability when compared to adjacent (in the chain) residues. Availability: http://biomine.ece.ualberta.ca/MoRFpred/; http://biomine.ece.ualberta.ca/MoRFpred/Supplement.pdf Contact: lkurgan@ece.ualberta.ca Supplementary information: Supplementary data are available at Bioinformatics online.

Do viral proteins possess unique biophysical features

Natural selection shapes the sequence, structure and biophysical properties of proteins to fit their environment. We hypothesize that highly thermostable proteins and viral proteins represent two opposing adaptation strategies. Thermostable proteins are highly compact and possess well-packed hydrophobic cores and intensely charged surfaces. By contrast, viral proteins, and RNA viral proteins in particular, display a high occurrence of disordered segments and loosely packed cores. These features might endow viral proteins with increased structural flexibility and effective ways to interact with the components of the host. They could also be related to high adaptability levels and mutation rates observed in viruses, thus, representing a unique strategy for buffering the deleterious effects of mutations, such that those that have little (interactions), have little to lose.