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We assessed whether in vivo mRNA structural reactivity and the structure of the encoded protein are related. This is the first investigation of such a relationship that utilizes information on RNA structure obtained in living cells. Based on our recent genome-wide Structure-seq analysis in Arabidopsis thaliana , we report that, as a meta property, regions of individual mRNAs that code for protein domains generally have higher reactivity to DMS (dimethyl sulfate), a chemical that covalently modifies accessible As and Cs, than regions that encode protein domain junctions. This relationship is prominent for proteins annotated for catalytic activity and reversed in proteins annotated for binding and transcription regulatory activity. Upon analyzing intrinsically disordered proteins, we found a similar pattern for disordered regions as compared to ordered regions: regions of individual mRNAs that code for ordered regions have significantly higher DMS reactivity than regions that code for intrinsically disordered regions. Based on these effects, we hypothesize that the decreased DMS reactivity of RNA regions that encode protein domain junctions or intrinsically disordered regions may reflect increased RNA structure that may slow translation, allowing time for the nascent protein domain or ordered region of the protein to fold, thereby reducing protein misfolding. In addition, a drop in DMS reactivity was observed on portions of mRNA sequences that correspond to the C-termini of protein domains, suggesting ribosome protection at these mRNA regions. Structural relationships between mRNAs and their encoded proteins may have evolved to allow efficient and accurate protein folding. [ABSTRACT FROM AUTHOR]

Abstract: We present a comprehensive bioinformatics analysis of the abundance and roles of intrinsic disorder in human proteins involved in the antiviral innate immune response. The commonness of intrinsic disorder and disorder-based binding sites is evaluated in 840 human antiviral proteins and proteins associated with innate immune response and defense response to virus. Among the mechanisms engaged in the innate immunity to viral infection are three receptor-based pathways activated by the specific recognition of various virus-associated patterns by several retinoic acid-inducible gene I-like receptors, toll-like receptors, and nucleotide oligomerization domain-like receptors. These modules are tightly regulated and intimately interconnected being jointly controlled via a complex set of protein–protein interactions. Focused analysis of the major players involved in these three pathways is performed to illustrate the roles of protein intrinsic disorder in controlling and regulating the innate antiviral immunity. We mapped the disorder into an integrated network of receptor-based pathways of human innate immunity to virus infection and demonstrate that proteins involved in regulation and execution of these innate immunity pathways possess substantial amount of intrinsic disorder. Disordered regions are engaged in a number of crucial functions, such as protein–protein interactions and interactions with other partners including nucleic acids and other ligands, and are enriched in posttranslational modification sites. Therefore, host cells use numerous advantages of intrinsically disordered proteins and regions to fight flexible invaders and viruses and to successfully overcome the viral invasion. [Copyright &y& Elsevier]

Abstract: Intrinsically disordered regions in proteins play active roles in recognition, signaling and molecular sorting. They often undergo coupled folding and binding giving rise to largely ordered interfaces with their binding partners. The cytoplasmic region of the T-cell receptor zeta subunit (ζcyt) has been previously proposed to specifically dimerize in the absence of a disorder-to-order transition, suggesting an intriguing dimerization mechanism that may involve multiple transient interfaces. We show here using analytical ultracentrifugation, NMR, size-exclusion chromatography (SEC) and multi-angle light scattering that neither ζcyt nor the cytoplasmic region of CD3ε significantly populates a dimeric state but that they are mostly monomers in solution up to millimolar concentrations. They experience a salt- and concentration-dependent shift of their elution volume in SEC previously interpreted as dimerization. Our data show that ζcyt does not form a highly disordered protein complex and leaves open the question as to whether completely disordered dimers (or other oligomers) exist in nature. [Copyright &y& Elsevier]

Abstract: Interdomain interactions of spectrin are critical for maintenance of the erythrocyte cytoskeleton. In particular, “head-to-head” dimerization occurs when the intrinsically disordered C-terminal tail of β-spectrin binds the N-terminal tail of α-spectrin, folding to form the “spectrin tetramer domain”. This non-covalent three-helix bundle domain is homologous in structure and sequence to previously studied spectrin domains. We find that this tetramer domain is surprisingly kinetically stable. Using a protein engineering Φ-value analysis to probe the mechanism of formation of this tetramer domain, we infer that the domain folds by the docking of the intrinsically disordered β-spectrin tail onto the more structured α-spectrin tail. [Copyright &y& Elsevier]

Abstract: In view of getting insights into the molecular determinants of the binding efficiency of intrinsically disordered proteins (IDPs), we used random mutagenesis. As a proof of concept, we chose the interaction between the intrinsically disordered C-terminal domain of the measles virus nucleoprotein (NTAIL) and the X domain (XD) of the viral phosphoprotein and assessed how amino acid substitutions introduced at random within NTAIL affect partner recognition. In contrast with directed evolution approaches, we did not apply any selection and used the gene library approach not for production purposes but for achieving a better understanding of the NTAIL/XD interaction. For that reason, and to differentiate our approach from similar approaches that make use of systematic (i.e., targeted) mutagenesis, we propose to call it “descriptive random mutagenesis” (DRM). NTAIL variants generated by error-prone PCR were picked at random in the absence of selection pressure and were characterized in terms of sequence and binding abilities toward XD. DRM not only identified determinants of NTAIL/XD interaction that were in good agreement with previous work but also provided new insights. In particular, we discovered that the primary interaction site is poorly evolvable in terms of binding abilities toward XD. We also identified a critical NTAIL residue whose role in stabilizing the NTAIL/XD complex had previously escaped detection, and we identified NTAIL regulatory sites that dampen the interaction while being located outside the primary interaction site. Results show that DRM is a valuable approach to study binding abilities of IDPs. [Copyright &y& Elsevier]

Abstract: Amyloid β (Aβ) peptides are a primary component of fibrils and oligomers implicated in the etiology of Alzheimer's disease (AD). However, the intrinsic flexibility of these peptides has frustrated efforts to investigate the secondary and tertiary structure of Aβ monomers, whose conformational landscapes directly contribute to the kinetics and thermodynamics of Aβ aggregation. In this work, de novo replica exchange molecular dynamics (REMD) simulations on the microseconds-per-replica timescale are used to characterize the structural ensembles of Aβ42, Aβ40, and M35-oxidized Aβ42, three physiologically relevant isoforms with substantially different aggregation properties. J-coupling data calculated from the REMD trajectories were compared to corresponding NMR-derived values acquired through two different pulse sequences, revealing that all simulations converge on the order of hundreds of nanoseconds-per-replica toward ensembles that yield good agreement with experiment. Though all three Aβ species adopt highly heterogeneous ensembles, these are considerably more structured compared to simulations on shorter timescales. Prominent in the C-terminus are antiparallel β-hairpins between L17–A21, A30–L36, and V39–I41, similar to oligomer and fibril intrapeptide models that expose these hydrophobic side chains to solvent and may serve as hotspots for self-association. Compared to reduced Aβ42, the absence of a second β-hairpin in Aβ40 and the sampling of alternate β topologies by M35-oxidized Aβ42 may explain the reduced aggregation rates of these forms. A persistent V24–K28 bend motif, observed in all three species, is stabilized by buried backbone to side-chain hydrogen bonds with D23 and a cross-region salt bridge between E22 and K28, highlighting the role of the familial AD-linked E22 and D23 residues in Aβ monomer folding. These characterizations help illustrate the conformational landscapes of Aβ monomers at atomic resolution and provide insight into the early stages of Aβ aggregation pathways. [Copyright &y& Elsevier]

Abstract: The poly-l-proline type II (PPII) helix in recent years has emerged clearly as a structural class not only of fibrillar proteins (in collagen, PPII is a dominant conformation) but also of the folded and unfolded proteins. Although much less abundant in folded proteins than the α-helix and β-structure, the left-handed, extended PPII helix represents the only frequently occurring regular structure apart from these two structure classes. Natively unfolded proteins have a high content of the PPII helices identified by spectroscopic methods. Apart from the structural function, PPII is favorable for protein–protein and protein–nucleic acid interactions and plays a major role in signal transduction and protein complex assembly, as this structure is often found in binding sites, specifically binding sites of widely spread SH3 domains. PPII helices do not necessarily contain proline, but proline has high PPII propensity. Commonly occurring proline-rich regions, serving as recognition sites, are likely to have PPII structure. PPII helices are involved in transcription, cell motility, self-assembly, elasticity, and bacterial and viral pathogenesis, and has an important structural role in amyloidogenic proteins. However, PPII helices are not always assigned in experimentally solved structures, and they are rarely used in protein structure modeling. We aim to give an overview of this structural class and of the place it holds in our current understanding of protein structure and function. This review is subdivided into three main parts: the first part covers PPII helices in unfolded peptides and proteins, the second part includes studies of the PPII helices in folded proteins, and the third part discusses the functional role of the PPII. [Copyright &y& Elsevier]