Scale-free networks and hubsIn biological systems, processes such as growth, energy generation, cell division and signaling are integrated by large, intricate networks. These biological networks, as well as certain nonbiological networks, especially those involved in communications such as the internet and cellular phone systems, are classified as scale-free networks (SFNs) [1][2][3]. The basic feature that separates these networks from non-SFNs such as regular Proteins participate in complex sets of interactions that represent the mechanistic foundation for much of the physiology and function of the cell. These protein-protein interactions are organized into exquisitely complex networks. The architecture of protein-protein interaction networks was recently proposed to be scale-free, with most of the proteins having only one or two connections but with relatively fewer 'hubs' possessing tens, hundreds or more links. The high level of hub connectivity must somehow be reflected in protein structure. What structural quality of hub proteins enables them to interact with large numbers of diverse targets? One possibility would be to employ binding regions that have the ability to bind multiple, structurally diverse partners. This trait can be imparted by the incorporation of intrinsic disorder in one or both partners. To illustrate the value of such contributions, this review examines the roles of intrinsic disorder in protein network architecture. We show that there are three general ways that intrinsic disorder can contribute: First, intrinsic disorder can serve as the structural basis for hub protein promiscuity; secondly, intrinsically disordered proteins can bind to structured hub proteins; and thirdly, intrinsic disorder can provide flexible linkers between functional domains with the linkers enabling mechanisms that facilitate binding diversity. An important research direction will be to determine what fraction of proteinprotein interaction in regulatory networks relies on intrinsic disorder. Abbreviations CaM, calmodulin; Cdk, cyclin-dependent protein kinase; CKI, Cdk inhibitor protein; GSK3b, glycogen synthase kinase 3 beta; ID, intrinsically disordered; MoRE, molecular recognition element; NER, nucleotide excision repair; PDB, Protein Data Bank; PONDRÒ, predictors of naturally disordered regions; RGN, regular network; RNN, random network; SFN, scale-free network; XPA, xeroderma pigmentosum group A protein; FRAT, frequently rearranged in advanced T-cell lymphomas; Wnt, wingless type MMTV integration site family; HMG, high mobility group; VL-XT, a predictor of intrinsic disorder that integrates various methods-based predictor of long disordered regions and X-ray based N-and Cterminal predictors; VSL1, length-dependent predictor of intrinsic protein disorder; RPA, replication protein A; ERCC1, exchange repair cross complementing complex 1; TFIIH, transcription factor IIH; XAB, XPA binding protein; p27Kip , cyclin-dependent kinase inhibitor protein p27/1B.

Intrinsically disordered proteins and regions carry out varied and vital cellular functions. Proteins with disordered regions are especially common in eukaryotic cells, with a subset of these proteins being mostly disordered, e.g., with more disordered than ordered residues. Two distinct methods have been previously described for using amino acid sequences to predict which proteins are likely to be mostly disordered. These methods are based on the net charge-hydropathy distribution and disorder prediction score distribution. Each of these methods is reexamined, and the prediction results are compared herein. A new prediction method based on consensus is described. Application of the consensus method to whole genomes reveals that approximately 4.5% of Yersinia pestis, 5% of Escherichia coli K12, 6% of Archaeoglobus fulgidus, 8% of Methanobacterium thermoautotrophicum, 23% of Arabidopsis thaliana, and 28% of Mus musculus proteins are mostly disordered. The unexpectedly high frequency of mostly disordered proteins in eukaryotes has important implications both for large-scale, high-throughput projects and also for focused experiments aimed at determination of protein structure and function.

The Database of Protein Disorder (DisProt) links structure and function information for intrinsically disordered proteins (IDPs). Intrinsically disordered proteins do not form a fixed three-dimensional structure under physiological conditions, either in their entireties or in segments or regions. We define IDP as a protein that contains at least one experimentally determined disordered region. Although lacking fixed structure, IDPs and regions carry out important biological functions, being typically involved in regulation, signaling and control. Such functions can involve high-specificity low-affinity interactions, the multiple binding of one protein to many partners and the multiple binding of many proteins to one partner. These three features are all enabled and enhanced by protein intrinsic disorder. One of the major hindrances in the study of IDPs has been the lack of organized information. DisProt was developed to enable IDP research by collecting and organizing knowledge regarding the experimental characterization and the functional associations of IDPs. In addition to being a unique source of biological information, DisProt opens doors for a plethora of bioinformatics studies. DisProt is openly available at .

Many protein-protein and protein-nucleic acid interactions involve coupled folding and binding of at least one of the partners. Here, we propose a protein structural element or feature that mediates the binding events of initially disordered regions. This element consists of a short region that undergoes coupled binding and folding within a longer region of disorder. We call these features "molecular recognition elements" (MoREs). Examples of MoREs bound to their partners can be found in the alpha-helix, beta-strand, polyproline II helix, or irregular secondary structure conformations, and in various mixtures of the four structural forms. Here we describe an algorithm that identifies regions having propensities to become alpha-helix-forming molecular recognition elements (alpha-MoREs) based on a discriminant function that indicates such regions while giving a low false-positive error rate on a large collection of structured proteins. Application of this algorithm to databases of genomics and functionally annotated proteins indicates that alpha-MoREs are likely to play important roles protein-protein interactions involved in signaling events.

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Alternative splicing of pre-mRNA generates two or more protein isoforms from a single gene, thereby contributing to protein diversity. Despite intensive efforts, an understanding of the protein structure-function implications of alternative splicing is still lacking. Intrinsic disorder, which is a lack of equilibrium 3D structure under physiological conditions, may provide this understanding. Intrinsic disorder is a common phenomenon, particularly in multicellular eukaryotes, and is responsible for important protein functions including regulation and signaling. We hypothesize that polypeptide segments affected by alternative splicing are most often intrinsically disordered such that alternative splicing enables functional and regulatory diversity while avoiding structural complications. We analyzed a set of 46 differentially spliced genes encoding experimentally characterized human proteins containing both structured and intrinsically disordered amino acid segments. We show that 81% of 75 alternatively spliced fragments in these proteins were associated with fully (57%) or partially (24%) disordered protein regions. Regions affected by alternative splicing were significantly biased toward encoding disordered residues, with a vanishingly small P value. A larger data set composed of 558 SwissProt proteins with known isoforms produced by 1,266 alternatively spliced fragments was characterized by applying the PONDR VSL1 disorder predictor. Results from prediction data are consistent with those obtained from experimental data, further supporting the proposed hypothesis. Associating alternative splicing with protein disorder enables the time-and tissue-specific modulation of protein function needed for cell differentiation and the evolution of multicellular organisms.evolution ͉ natively unfolded ͉ intrinsically unstructured ͉ protein structure T he splicing of pre-mRNA (1) was first described in 1977. Soon thereafter, Gilbert (2) coined the terms ''intron'' (intragenic region) and ''exon'' (expressed region) for the noncoding and coding regions, respectively. Alternative splicing occurs when different mRNAs are assembled from a single gene by joining exons in different ways. Alternative splicing is proposed to generate complexity in multicellular eukaryotes by increasing protein diversity, and thus proteome size, from a relatively small number of genes (3). Estimates indicate that between 35 and 60% of human genes yield protein isoforms by means of alternatively spliced (AS) mRNA (4). Furthermore, complexity in higher organisms is also brought about by signaling and regulatory networks that enable robustness (5). The importance of alternative splicing as a regulatory process (3, 6) has been highlighted by the high occurrence of such splicing in the pre-mRNAs of regulatory and signaling proteins (7).Alternative splicing can bolster organism complexity, not only by effectively increasing proteome size and regulatory and signaling network complexity, but also by doing so in a time-and tissue-specific manner, supporting ...

Molecular Recognition Features (MoRFs) are short, interaction-prone segments of protein disorder that undergo disorder-to-order transitions upon specific binding, representing a specific class of intrinsically disordered regions that exhibit molecular recognition and binding functions. MoRFs are common in various proteomes and occupy a unique structural and functional niche in which function is a direct consequence of intrinsic disorder. Example MoRFs collected from the Protein Data Bank (PDB) have been divided into three subtypes according to their structures in the bound state: α-MoRFs form α-helices, β-MoRFs form β-strands, and ι-MoRFs form structures without a regular pattern of backbone hydrogen bonds. These example MoRFs were indicated to be intrinsically disordered in the absence of their binding partners by several criteria. In this study we used several geometric and physiochemical criteria to examine the properties of 62 α-, 20 β-and 176 ι-MoRF complex structures. Interface residues were examined by calculating differences in accessible surface area between the complex and isolated monomers. The compositions and physiochemical properties of MoRF and MoRF partner interface residues were compared to the interface residues of homodimers, heterodimers, and antigen-antibody complexes. Our analysis indicates that there are significant differences in residue composition and several geometric and physicochemical properties that can be used to discriminate, with a high degree of accuracy, between various interfaces in protein interaction datasets. Implications of these findings for the development of MoRF-partner interaction predictors are discussed. In addition, structural changes upon MoRF-to-partner complex formation were examined for several illustrative examples. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptProtein-protein interaction sites have been intensively analyzed by different researchers to understand the molecular determinants of protein recognition and to identify specific characteristics of protein-protein interfaces. 1-18 Different aspects of interaction sites, including residue propensities, residue pairing preferences, hydrophobicity, size, shape, solvent accessibility, and hydrogen bond protection, have all been examined. Although each of these parameters provides some information indicative of protein-protein interaction sites, none of them perfectly differentiates interaction sites from noninteracting protein surfaces. Protein interaction sites have been observed to be hydrophobic, planar, globular and protruding. 1, 2, 4, 8, 9, 16 Furthermore, interfaces in different types of even the simplest protein complexes (e.g., homodimers, heterodimers) have different properties 9, 11, 15 . Homocomplexes are often permanent and optimized, whereas many heterocomplexes are nonobligatory, associating and disassociating according to the environmental or external factors and involve proteins that must also exist independently. 9 Subunit interfaces in stable oligomer...

Regulation, recognition and cell signaling involve the coordinated actions of many players. Signaling scaffolds, with their ability to bring together proteins belonging to common and/or interlinked pathways, play crucial roles in orchestrating numerous events by coordinating specific interactions among signaling proteins. This review examines the roles of intrinsic disorder (ID) in signaling scaffold protein function. Several well-characterized scaffold proteins with structurally and functionally characterized ID regions are used here to illustrate the importance of ID for scaffolding function. These examples include scaffolds that are mostly disordered, only partially disordered or those in which the ID resides in a scaffold partner. Specific scaffolds discussed include RNase, voltage-activated potassium channels, axin, BRCA1, GSK-3beta, p53, Ste5, titin, Fus3, BRCA1, MAP2, D-AKAP2 and AKAP250. Among the mechanisms discussed are: molecular recognition features, fly-casting, ease of encounter complex formation, structural isolation of partners, modulation of interactions between bound partners, masking of intramolecular interaction sites, maximized interaction surface per residue, toleration of high evolutionary rates, binding site overlap, allosteric modification, palindromic binding, reduced constraints for alternative splicing, efficient regulation via posttranslational modification, efficient regulation via rapid degradation, protection of normally solvent-exposed sites, enhancing the plasticity of interaction and molecular crowding. We conclude that ID can enhance scaffold function by a diverse array of mechanisms. In other words, scaffold proteins utilize several ID-facilitated mechanisms to enhance function, and by doing so, get more functionality from less structure.