Biology subjects

Milanetti, E.

Publications and source records attributed to Milanetti, E..

11 recordsLinked to original sources

bioRxiv2026

Microsecond molecular dynamics of SOD1 variants suggest a structural basis for divergent ALS clinical outcomes

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterised by progressive motor neuron degeneration. Mutations in the SOD1 gene represent the second most common genetic cause of ALS (ALS), and distinct SOD1 missense variants present with markedly different clinical profiles. A4V leads to an aggressive form of the disease (median survival [~]1y), H46R confers a mild, slowly progressive course and I113T exhibits an intermediate phenotype. The molecular basis by which these mutations produce divergent clinical outcomes remains poorly understood. We performed extensive classical molecular dynamics simulations of wild-type SOD1 and the three ALS-associated variants in the apo monomeric state to attempt to investigate the mechanisms behind such phenotypic differences. Structural stability, global compactness, and conformational flexibility, as well as analysis of collective motions between residues and estimation of free energy, were assessed. The H46R, A4V, and I113T variants exhibited distinct dynamic behaviours, highlighting differences in structural stability, local flexibility, and intramolecular interactions. These findings suggest that specific structural regions may contribute differently to protein dysfunction and could represent key elements for understanding the relationship between molecular dynamic properties and the differing clinical severity associated with these variants. Most strikingly, H46R exhibited exceptional structural stability across every analytical level, the lowest global deviation, most attenuated local flexibility, strongest internal dynamic coordination, and the deepest, most confined free energy basins of any system examined. This convergent multi-layered evidence of structural restraint provides a compelling mechanistic basis for the mild and slowly progressive clinical course of H46R ALS, suggesting that enhanced conformational rigidity, rather than bulk destabilisation, is the defining biophysical feature of this variant, and that its pathogenic mechanism operates through a route fundamentally decoupled from the aggregation-driven toxicity that characterises the more aggressive SOD1-ALS mutations.

genomics↗

bioRxiv2026

α-Synuclein and γ-Tubulin Cooperatively Regulate Activity-Evoked Presynaptic Microtubule Nucleation to Gate Dopamine Release

-Synuclein has long been implicated in the regulation of synaptic activity, but the molecular basis that underlies this function has been elusive. Here, we identify a microtubule (MT)-dependent mechanism through which -synuclein regulates synaptic dopamine release. Using live imaging of cultured dopaminergic neurons, we visualize dynamic MTs at individual presynaptic boutons and show that neuronal activity triggers local {gamma}-tubulin-dependent MT nucleation. We find that this nucleation is essential for interbouton synaptic vesicle (SV) transport and for sustained dopamine release during high activity. We further discover that -synuclein acts as a positive regulator of presynaptic MT nucleation by binding directly to {gamma}-tubulin and the /{beta}-tubulin heterodimer. Activity-evoked phosphorylation of -synuclein at serine 129, a modification that accumulates in synucleinopathies and a molecular switch for -synuclein binding to synaptic proteins, occurs in the region of /m tubulin binding and is both necessary and sufficient for MT initiation. Our findings reveal a previously unrecognized, activity-dependent role for -synuclein in the nucleation of axonal MTs that enables on-demand SV interbouton redistribution and dopamine release. This mechanism provides a novel molecular link between -synuclein phosphorylation and MT-dependent modulation of dopamine release, offering insight into how its dysregulation may contribute to dopaminergic synaptic dysfunction, a central feature of synucleinopathies.

cell biology↗

bioRxiv2025

Preventing pathogenic dimerization in a misfolded antibody light chain through the design of an inhibitory peptide

Immunoglobulin light chain (AL) amyloidosis is the most common form of systemic amyloidosis. The disease correlates with the formation of insoluble aggregates mostly composed by patient-specific antibody light chains, whose hypervariable regions make each case unique and highlight the need for personalized therapeutics. In this study, we focused on a pathogenic homodimer we previously obtained from a patient-derived light chain. By analyzing the dynamics and the interface of this dimer, we identified a 15-residue peptide with potential inhibitory activity. The peptide was then refined using a computational mutagenesis protocol that iteratively improved its sequence to maximize complementarity with the protein interface, taking into account shape, electrostatics, and hydropathy. The resulting optimized peptide is found to bind the monomer with a binding affinity comparable to that of the full pathogenic interface. These results suggest that the designed peptide could act as an effective antagonist of the pathogenic dimer, and demonstrate that our computational strategy could provide a general framework for designing patient-specific inhibitory peptides against aggregation-prone proteins.

biochemistry↗

bioRxiv2024

Towards universal models for collective interactions in biomolecular condensates

A wide-range of higher-order structures, including dense, liquid-like assemblies, serve as key components of cellular matter. The molecular language of how protein sequences encode the formation and biophysical properties of biomolecular condensates, however, is not completely understood. Recent notion on the scale invariance of the cluster sizes below the critical concentration for phase separation suggests a universal mechanism, which can operate from oligomers to non-stoichiometric assemblies. Here we propose a model for collective interactions in condensates, based on context-dependent, variable interactions. We provide the mathematical formalism, which is capable describing growing dynamic clusters as well as changes in their material properties. Furthermore, we discuss the consequences of the model to maximise sensitivity to the environmental signals and to increase correlation lengths.

systems biology↗

bioRxiv2022

Evolutionary dynamics of genome size and content during the adaptive radiation of Heliconiini butterflies

Heliconius butterflies, a speciose genus of Mullerian mimics, represent a classic example of an adaptive radiation that includes a range of derived dietary, life history, physiological and neural traits. However, key lineages within the genus, and across the broader Heliconiini tribe, lack genomic resources, limiting our understanding of how adaptive and neutral processes shaped genome evolution during their radiation. We have generated highly contiguous genome assemblies for nine new Heliconiini, 29 additional reference-assembled genomes, and improve 10 existing assemblies. Altogether, we provide a major new dataset of annotated genomes for a total of 63 species, including 58 species within the Heliconiini tribe. We use this extensive dataset to generate a robust and dated heliconiine phylogeny, describe major patterns of introgression, explore the evolution of genome architecture, and the genomic basis of key innovations in this enigmatic group, including an assessment of the evolution of putative regulatory regions at the Heliconius stem. Our work illustrates how the increased resolution provided by such dense genomic sampling improves our power to generate and test gene-phenotype hypotheses, and precisely characterize how genomes evolve.

evolutionary biology↗

bioRxiv2022

A role of Lck annular lipids in the steady upkeep of active Lck in T cells

Theoretical work suggests that collective spatiotemporal behaviour of integral membrane proteins (IMPs) can be modulated by annular lipids sheathing their hydrophobic moiety. Here, we present evidence for this prediction in a natural membrane by investigating the mechanism that maintains steady amount of active isoform of Lck kinase (LckA) by Lck trans-autophosphorylation offset by the phosphatase CD45. We gauged experimental suitability by quantitation of CD45 and LckA subcellular localisation, LckA generation as a function of Lck and pharmacological perturbation. Steady LckA was challenged by swapping Lck membrane anchor with structurally divergent ones expected to substantially modify Lck annular lipids, such as that of Src or the transmembrane domains of LAT, CD4, palmitoylation-defective CD4 and CD45, respectively. The data showed only small alteration of LckA, except for CD45 hydrophobic anchor that thwarted LckA, due to excessive lateral proximity to CD45. The data are best explained by annular lipids facilitating or penalising IMPs lateral proximity, hence modulating IMPs protein-protein functional interactions. Our findings can contribute to improve the understanding of biomembranes organisation.

cell biology↗

bioRxiv2021

Inferring the stabilization effects of SARS-CoV-2 variants on the binding with ACE2 receptor

With the progression of the SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) pandemic, several variants of the virus are emerging with mutations distributed all over the viral sequence. While most of them are expected to have little to no effects at the phenotype level, some of these variants presenting specific mutations on the Spike protein are rapidly spreading, making urgent the need of characterizing their effects on phenotype features like contagiousness and antigenicity. With this aim, we performed extensive molecular dynamics simulations on a selected set of possible Spike variants in order to assess the stabilizing effect of particular amino acid substitutions, with a special focus on the mutations that are both characteristic of the top three most worrying variants at the moment, i.e the English, South African and Amazonian ones, and that occur at the molecular interface between SARS-CoV-2 Spike protein and its human ACE2 receptor. We characterize these variants effect in terms of (i) residues mobility, (ii) compactness, studying the network of interactions at the interface, and (iii) variation of shape complementarity via expanding the molecular surfaces in the Zernike basis. Overall, our analyses highlighted greater stability of the three variant complexes with respect to both the wild type and two negative control systems, especially for the English and Amazonian variants. In addition, in the three variants, we investigate the effects a not-yet observed mutation in position 501 could provoke on complex stability. We found that a phenylalanine mutation behaves similarly to the English variant and may cooperate in further increasing the stability of the South African one, hinting at the need for careful surveillance for the emergence of such kind of mutations in the population. Ultimately, we show that the observables we propose describe key features for the stability of the ACE2-spike complex and can help to monitor further possible spike variants.

biophysics↗

bioRxiv2020

Structural analysis of SARS-CoV-2 andprediction of the human interactome

Specific elements of viral genomes regulate interactions within host cells. Here, we calculated the secondary structure content of >2000 coronaviruses and computed >100000 human protein interactions with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The genomic regions display different degrees of conservation. SARS-CoV-2 domain encompassing nucleotides 22500 - 23000 is conserved both at the sequence and structural level. The regions upstream and downstream, however, vary significantly. This part codes for the Spike S protein that interacts with the human receptor angiotensin-converting enzyme 2 (ACE2). Thus, variability of Spike S may be connected to different levels of viral entry in human cells within the population. Our predictions indicate that the 5 end of SARS-CoV-2 is highly structured and interacts with several human proteins. The binding proteins are involved in viral RNA processing such as double-stranded RNA specific editases and ATP-dependent RNA-helicases and have strong propensity to form stress granules and phase-separated complexes. We propose that these proteins, also implicated in viral infections such as HIV, are selectively recruited by SARS-CoV-2 genome to alter transcriptional and post-transcriptional regulation of host cells and to promote viral replication.

bioinformatics↗

bioRxiv2020

In-Silico evidence for two receptors based strategy of SARS-CoV-2

We propose a novel numerical method able to determine efficiently and effectively the relationship of complementarity between portions of protein surfaces. This innovative and general procedure, based on the representation of the molecular iso-electron density surface in terms of 2D Zernike polynomials, allows the rapid and quantitative assessment of the geometrical shape complementarity between interacting proteins, that was unfeasible with previous methods. We first tested the method with a large dataset of known protein complexes obtaining an overall area under the ROC curve of 0.76 in the blind recognition of binding sites and then applied it to investigate the features of the interaction between the Spike protein of SARS-CoV-2 and human cellular receptors. Our results indicate that SARS-CoV-2 uses a dual strategy: its spike protein could also interact with sialic acid receptors of the cells in the upper airways, in addition to the known interaction with Angiotensin-converting enzyme 2.

biophysics↗

bioRxiv2020

proABC-2: PRediction Of AntiBody Contacts v2 and its application to information-driven docking

Monoclonal antibodies (mAbs) are essential tools in the contemporary therapeutic armoury. Understanding how these recognize their antigen is a fundamental step in their rational design and engineering. The rising amount of publicly available data is catalysing the development of computational approaches able to offer valuable, faster and cheaper alternatives to classical experimental methodologies used for the study of antibody-antigen complexes. Here we present proABC-2, an update of the original random-forest antibody paratope predictor, based on a convolutional neural network algorithm. We also demonstrate how the predictions can be fruitfully used to drive the docking in HADDOCK. The proABC-2 server is freely available at: https://bianca.science.uu.nl/proabc2/.

bioinformatics↗

bioRxiv2018

Insights on protein thermal stability: a graph representation of molecular interactions

Understanding the molecular mechanisms of thermal stability is a challenge in protein biology. Indeed, knowing the temperature at which proteins are stable has important theoretical implications, which are intimately linked with properties of the native fold, and a wide range of potential applications from drug design to the optimization of enzyme activity.\n\nHere, we present a novel graph-theoretical framework to assess thermal stability based on the structure without any a priori information. In our approach we describe proteins as energy-weighted graphs and compare them using ensembles of interaction networks. Investigating the position of specific interactions within the 3D native structure, we developed a parameter-free network descriptor that permits to distinguish thermostable and mesostable proteins with an accuracy of 76% and Area Under the Roc Curve of 78%.

bioinformatics↗