Biology subjects

Potestio, R.

Publications and source records attributed to Potestio, R..

6 recordsLinked to original sources

bioRxiv2022

Kinetics of radiation-induced DNA double-strand breaks through coarse-grained simulations

Double-strand breaks (DSBs), i.e. the covalent cut of the DNA backbone over both strands, are a detrimental outcome of cell irradiation, bearing chromosomal aberrations and leading to cell apoptosis. In the early stages of the evolution of a DSB, the disruption of the residual interactions between the DNA moieties drives the fracture of the helical layout; in spite of its biological significance, the details of this process are still largely uncertain. Here, we address the mechanical rupture of DNA by DSBs via coarse-grained molecular dynamics simulations: the setup involves a 3855-bp DNA filament and diverse DSB motifs, i.e. within a range of distances between strand breaks (or DSB distance). By employing a coarse-grained model of DNA, we access the molecular details and characteristic timescales of the rupturing process. A sequence-nonspecific, linear correlation is observed between the DSB distance and the internal energy contribution to the disruption of the residual (Watson-Crick and stacking) contacts between DNA moieties, which is seemingly driven by an abrupt, cooperative process. Moreover, we infer an exponential dependence of the characteristic rupture times on the DSB distances, which we associate to an Arrhenius law of thermally-activated processes. This work lays the foundations of a detailed, mechanistic assessment of DSBs in silico, as a benchmark to both numerical simulations and data from single molecule experiments.

biophysics↗

bioRxiv2022

In search of a dynamical vocabulary: a pipeline to construct a basis of shared traits in large-scale motions of proteins

The paradigmatic sequence-structure-dynamics-function relation in proteins is nowadays well established in the scientific community; in particular, large effort has been spent to probe the first connection, indeed providing convincing evidence of its strength and rationalising it in a quantitative and general framework. In contrast, however, the role of dynamics as a link between structure and function has eluded a similarly clear-cut verification and description. In this work, we propose a pipeline aimed at building a basis for the quantitative characterisation of the large-scale dynamics of a set of proteins, starting from the sole knowledge of their native structures. The method hinges on a dynamics-based clusterization, which allows a straightforward comparison with structural and functional protein classifications. The resulting basis set, obtained through the application to a group of related proteins, is shown to reproduce the salient large-scale dynamical features of the dataset. Most interestingly, the basis set is shown to encode the fluctuation patterns of homologous proteins not belonging to the initial dataset, thus highlighting the general applicability of the pipeline used to build it.

biophysics↗

bioRxiv2022

Structural Basis of Mutation-Dependent p53 Tetramerization Deficiency

The formation of a tetrameric assembly is essential for the ability of the tumor suppressor protein p53 to act as a transcription factor. Such a quaternary conformation is driven by a specific tetramerization domain, separated from the central DNA binding domain by a flexible linker. Despite the distance, functional crosstalk between the two domains has been reported. This phenomenon can explain the pathogenicity of some inherited or somatically acquired mutations in the tetramerization domain, including the widespread R337H missense mutation occurring in the population of south Brazil. In this work, we have combined computational predictions through extended all-atom molecular dynamics simulations with functional assays in a genetically defined yeast-based model system to reveal structural features of p53 tetramerization domains and their transactivation capacity and specificity. Besides the germline and cancer-associated R337H and R337C, other rationally designed missense mutations targeting a significant salt bridge interaction that stabilizes the p53 tetramerization domain were studied (R337D, D352R, and the double mutation R337D plus D352R). Simulations revealed a destabilizing effect of pathogenic mutations within the p53 tetramerization domain and highlighted the importance of electrostatic interactions between residues 337 and 352. The transactivation assay performed in yeast by tuning the expression of wild-type and mutant p53 proteins revealed that p53 tetramerization mutations could decrease transactivation potential and alter transactivation specificity, in particular, by better tolerating the negative features in weak DNA binding sites. These results establish the effect of naturally occurring variations at positions 337 and 352 on p53 conformational stability and function.

cancer biology↗

bioRxiv2022

Membrane binding of pore-forming γ-hemolysin components studied at different lipid compositions

Methicillin-resistant Staphylococcus aureus is is among those pathogens currently posing the highest threat to public health. Its host immune evasion strategy is mediated by pore-forming toxins (PFTs), among which the bicomponent {gamma}-hemolysin is one of the most common. The complexity of the porogenesis mechanism by {gamma}-hemolysin poses difficulties in the development of antivirulence therapies targeting PFTs from S. aureus, and sparse and apparently contrasting experimental data have been produced. Here, through a large set of molecular dynamics simulations at different levels of resolution, we investigate the first step of pore formation, and in particular the effect of membrane composition on the ability of{gamma} -hemolysin components, LukF and Hlg2, to steadily adhere to the lipid bilayer in the absence of proteinaceous receptors. Our simulations are in agreement with experimental data of {gamma}-hemolysin pore formation on model membranes, which are here explained on the basis of the bilayer properties. Our computational investigation suggests a possible rationale to explain experimental data on phospholipid binding to the LukF component, and to hypothesise a mechanism by which, on purely lipidic bilayers, the stable anchoring of LukF to the cell surface facilitates Hlg2 binding, through the exposure of its N-terminal region. We expect that further insights on the mechanism of transition between soluble and membrane bound-forms and on the role played by the lipid molecules will contribute to the design of antivirulence agents with enhanced efficacy against methicillin-resistant S. aureus infections. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/479512v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@5f0ae5org.highwire.dtl.DTLVardef@1c46688org.highwire.dtl.DTLVardef@1277e17org.highwire.dtl.DTLVardef@1c71898_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIThe presence of cholesterol and unsaturated phospholipid tails facilitates the binding of{gamma} -hemolysin components, LukF and Hlg2, on model membranes. C_LIO_LICoarse-grained simulations show that the two components have different absorption capabilities, with LukF undergoing the most stable binding. C_LIO_LIThe spontaneous docking of LukF on the membrane is mediated by two distant phosphatidylcholine binding sites. C_LI

biophysics↗

bioRxiv2021

How Communication Pathways Bridge Local and Global Conformations in an IgG4 Antibody: a Molecular Dynamics Study

The affinity of an antibody for its antigen is primarily determined by the specific sequence and structural arrangement of the complementarity-determining regions (CDRs). Recently, however, evidence has accumulated that points toward a nontrivial relation between the CDR and distal sites on the antibody structure: variations in the binding strengths have been observed upon mutating amino acids separated from the paratope by several nanometers, thus suggesting the existence of a communication network within antibodies whose extension and relevance might be deeper than insofar expected. In this work, we test this hypothesis by means of molecular dynamics (MD) simulations of the IgG4 monoclonal antibody pembrolizumab, an approved drug that targets the programmed cell death protein 1 (PD-1). The molecule is simulated in both the apo and holo states, totalling 4s of MD trajectory. The analysis of these simulations shows that the bound antibody explores a restricted range of conformations with respect to the apo one, and that the global conformation of the molecule correlates with that of the CDR; a pivotal role in this relationship is played by the relatively short hinge, which mechanically couples Fab and Fc domains. These results support the hypothesis that pembrolizumab behaves as a complex machinery, with a multi-scale hierarchy of global and local conformational changes that communicate with one another. The analysis pipeline developed in this work is general, and it can help shed further light on the mechanistic aspects of antibody function. SynopsisAntigen binding restricts the conformational variability of the therapeutic antibody pembrolizumab in an interplay between the paratope and hinge region, mediated by a full-scale interaction network. Graphical TOC Entry O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/449604v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@d64d7corg.highwire.dtl.DTLVardef@17919c2org.highwire.dtl.DTLVardef@18a21bforg.highwire.dtl.DTLVardef@1adb5a7_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2018

Searching the optimal folding routes of a Complex Lasso protein

Understanding how polypeptides can efficiently and reproducibly attain a self-entangled conformation is a compelling biophysical challenge, which might shed new light on our general knowledge of protein folding. Complex Lassos, namely self-entangled protein structures characterized by a covalent loop sealed by a cysteine bridge, represent an ideal test system in the framework of entangled folding. Indeed, as cysteine bridges form in oxidizing conditions, they can be used as on/off switches of the structure topology, to investigate the role played by the backbone entanglement in the process. In the present work we have used molecular dynamics to simulate the folding of a complex lasso glycoprotein, Granulocyte-macrophage colony-stimulating factor, modeling both reducing and oxidizing conditions. Together with a well-established Go-like description, we have employed the elastic folder model, a Coarse-Grained, minimalistic representation of the polypeptide chain, driven by a structure-based angular potential. The purpose of this study is to assess the kinetically optimal pathways, in relation to the formation of the native topology. To this end we have implemented an evolutionary strategy that tunes the elastic folder model potentials to maximize the folding probability within the early stages of the dynamics. The resulting protein model is capable of folding with high success rate, avoiding the kinetic traps that hamper the efficient folding in the other tested models. Employing specifically designed topological descriptors, we could observe that the selected folding routes avoid the topological bottleneck by locking the cysteine bridge after the topology is formed. These results provide valuable insights on the selection of mechanisms in self-entangled protein folding while, at the same time, the proposed methodology can complement the usage of established minimalistic models, and draw useful guidelines for more detailed simulations.

biophysics↗