Computational approaches for inferring the functions of intrinsically disordered proteins.

Intrinsically disordered proteins (IDPs) are ubiquitously involved in cellular processes and often implicated in human pathological conditions. The critical biological roles of these proteins, despite not adopting a well-defined fold, encouraged structural biologists to revisit their views on the protein structure-function paradigm. Unfortunately, investigating the characteristics and describing the structural behavior of IDPs is far from trivial, and inferring the function(s) of a disordered protein region remains a major challenge. Computational methods have proven particularly relevant for studying IDPs: on the sequence level their dependence on distinct characteristics determined by the local amino acid context makes sequence-based prediction algorithms viable and reliable tools for large scale analyses, while on the structure level the in silico integration of fundamentally different experimental data types is essential to describe the behavior of a flexible protein chain. Here, we offer an overview of the latest developments and computational techniques that aim to uncover how protein function is connected to intrinsic disorder

DisCons: a novel tool to quantify and classify evolutionary conservation of intrinsic protein disorder

BACKGROUND: Analyzing the amino acid sequence of an intrinsically disordered protein (IDP) in an evolutionary context can yield novel insights on the functional role of disordered regions and sequence element(s). However, in the case of many IDPs, the lack of evolutionary conservation of the primary sequence can hamper the study of functionality, because the conservation of their disorder profile and ensuing function(s) may not appear in a traditional analysis of the evolutionary history of the protein. RESULTS: Here we present DisCons (Disorder Conservation), a novel pipelined tool that combines the quantification of sequence- and disorder conservation to classify disordered residue positions. According to this scheme, the most interesting categories (for functional purposes) are constrained disordered residues and flexible disordered residues. The former residues show conservation of both the sequence and the property of disorder and are associated mainly with specific binding functionalities (e.g., short, linear motifs, SLiMs), whereas the latter class correspond to segments where disorder as a feature is important for function as opposed to the identity of the underlying sequence (e.g., entropic chains and linkers). DisCons therefore helps with elucidating the function(s) arising from the disordered state by analyzing individual proteins as well as large-scale proteomics datasets. CONCLUSIONS: DisCons is an openly accessible sequence analysis tool that identifies and highlights structurally disordered segments of proteins where the conformational flexibility is conserved across homologs, and therefore potentially functional. The tool is freely available both as a web application and as stand-alone source code hosted at http://pedb.vib.be/discons

Biotemplated synthesis of magnetic filaments

With the aim of creating one-dimensional magnetic nanostructures, we genetically engineered flagellar filaments produced by Salmonella bacteria to display iron- or magnetite-binding sites, and used the mutant filaments as templates for both nucleation and attachment of the magnetic iron oxide magnetite.</p

Affinity purification of human m-calpain through an intrinsically disordered inhibitor, calpastatin

Calpains are calcium-activated proteases that have biomedical and biotechnological potential. Their activity is tightly regulated by their endogenous inhibitor, calpastatin that binds to the enzyme only in the presence of calcium. Conventional approaches to purify calpain comprise multiple chromatographic steps, and are labor-intensive, leading to low yields. Here we report a new purification procedure for the human m-calpain based on its reversible calcium-mediated interaction with the intrinsically disordered calpastatin. We exploit the specific binding properties of human calpastatin domain 1 (hCSD1) to physically capture human m-calpain from a complex biological mixture. The dissociation of the complex is mediated by chelating calcium, upon which heterodimeric calpain elutes while hCSD1 remains immobilized onto the stationary phase. This novel affinity-based purification was compared to the conventional multistep purification strategy and we find that it is robust, it yields a homogeneous preparation, it can be scaled up easily and it rests on a non-disruptive step that maintains close to physiological conditions that allow further biophysical and functional studies. © 2017 Nguyen et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

Distribution and composition of Mg-calcite and dolomite in the water and sediments of Lake Balaton

Lake Balaton is a large and shallow lake that is of great economic and cultural importance in landlocked Hungary. Even though the lake has been studied extensively in the last century from a large number of scientific aspects, the mineralogy of its sediments has not been fully explored. The mud at the bottom of the lake consists mostly of silt-sized grains of carbonate minerals with compositions between those of calcite (CaCO3) and dolomite CaMg (CO3)2. In order to understand the processes of carbonate precipitation and the influence of water budget fluctuations on the mineralogical character of the sediment, we used X-ray powder diffraction to analyze the changes of cell parameters of carbonate minerals in the upper half meter of the sediment. The major carbonate phase is Mg-calcite that shows a distinct reduction in cell parameters from west to east, reflecting an increase of its Mg-content, in parallel with a gradient of dissolved Mg/Ca ratio in the water. Intriguingly, dolomite, the other widespread carbonate phase in the sediment, also shows a change in cell parameters from west to east, with the deviations from values of stoichiometric dolomite being largest in the Eastern Basin of the lake. The similar pattern of cell parameter changes of Mg-calcite and dolomite suggests that ordered dolomite with slightly anomalous, Ca-rich composition also forms in the lake, probably by direct precipitation from the water. In contrast, protodolomite forms within the sediment through diagenetic processes. Based on our X-ray powder diffraction measurements, we propose a model of carbonate mineral formation and transformation in Lake Balaton. Since the Mg/Ca ratio of the water appears to be the major factor in controlling the compositions of carbonate minerals, and this ratio in turn is governed by the amount of water supply, the properties of the precipitating carbonate minerals are affected by the actual level of the lake water

Start2Fold: a database of hydrogen/deuterium exchange data on protein folding and stability

Proteins fulfil a wide range of tasks in cells; understanding how they fold into complex three-dimensional (3D) structures and how these structures remain stable while retaining sufficient dynamics for functionality is essential for the interpretation of overall protein behaviour. Since the 1950's, solvent exchange-based methods have been the most powerful experimental means to obtain information on the folding and stability of proteins. Considerable expertise and care were required to obtain the resulting datasets, which, despite their importance and intrinsic value, have never been collected, curated and classified. Start2Fold is an openly accessible database (http://start2fold.eu) of carefully curated hydrogen/deuterium exchange (HDX) data extracted from the literature that is open for new submissions from the community. The database entries contain (i) information on the proteins investigated and the underlying experimental procedures and (ii) the classification of the residues based on their exchange protection levels, also allowing for the instant visualization of the relevant residue groups on the 3D structures of the corresponding proteins. By providing a clear hierarchical framework for the easy sharing, comparison and (re-)interpretation of HDX data, Start2Fold intends to promote a better understanding of how the protein sequence encodes folding and structure as well as the development of new computational methods predicting protein folding and stability

AmyPro: a database of proteins with validated amyloidogenic regions

Soluble functional proteins may transform into insoluble amyloid fibrils that deposit in a variety of tissues. Amyloid formation is a hallmark of age-related degenerative disorders. Perhaps surprisingly, amyloid fibrils can also be beneficial and are frequently exploited for diverse functional roles in organisms. Here we introduce AmyPro, an open-access database providing a comprehensive, carefully curated collection of validated amyloid fibril-forming proteins from all kingdoms of life classified into broad functional categories (http://amypro.net). In particular, AmyPro provides the boundaries of experimentally validated amyloidogenic sequence regions, short descriptions of the functional relevance of the proteins and their amyloid state, a list of the experimental techniques applied to study the amyloid state, important structural/functional/variation/mutation data transferred from UniProt, a list of relevant PDB structures categorized according to protein states, database cross-references and literature references. AmyPro greatly improves on similar currently available resources by incorporating both prions and functional amyloids in addition to pathogenic amyloids, and allows users to screen their sequences against the entire collection of validated amyloidogenic sequence fragments. By enabling further elucidation of the sequential determinants of amyloid fibril formation, we hope AmyPro will enhance the development of new methods for the precise prediction of amyloidogenic regions within proteins

Linking functions: an additional role for an intrinsically disordered linker domain in the transcriptional coactivator CBP

The multi-domain transcriptional coactivators CBP/p300 integrate a multitude of signaling inputs, interacting with more than 400 proteins via one or more of their globular domains. While CBP/p300 function is typically considered in terms of these structured domains, about half of the protein consists of intrinsically disordered regions (IDRs) of varying length. However, these IDRs have only been thought of as linkers that allow flexible spatial arrangement of the structured domains, but recent studies have shown that similar IDRs mediate specific and critical interactions in other proteins. To examine the roles of IDRs in CBP, we performed yeast-two-hybrid screenings of placenta and lung cancer cDNA libraries, which demonstrated that the long IDR linking the KIX domain and bromodomain of CBP (termed ID3) can potentially bind to several proteins. The RNA-binding Zinc-finger protein 106 (ZFP106) detected in both libraries was identified as a novel substrate for CBP-mediated acetylation. Nuclear magnetic resonance (NMR) spectroscopy combined with cross-linking experiments and competition-binding assays showed that the fully disordered isolated ID3 transiently interacts with an IDR of ZFP106 in a fashion that disorder of both regions is maintained. These findings demonstrate that beside the linking function, ID3 can also interact with acetylation substrates of CBP