Biology subjects

Lovas, S.

Publications and source records attributed to Lovas, S..

7 recordsLinked to original sources

bioRxiv2026

Laminin-Derived Peptides Bind Neuropilin-1 at the VEGF-A165 Binding Pocket

Neuropilin-1 (NRP-1) is associated with glioblastoma progression and contains a b1-domain pocket that binds the C-terminal region of VEGF-A165. Whether laminin-derived peptide fragments can associate with this site is unknown. AlphaFold 3 complex prediction followed by 15 s of all-atom production molecular dynamics simulation, trajectory-based pocket occupancy and interaction analyses, and FoldX energetic scoring was performed to compare binding of the laminin-derived pentapeptide Ac-Tyr-Ile-Gly-Ser-Arg-NH2 (peptide 1) with its retro (Ac-Arg-Ser-Gly-Ile-Tyr-NH2, peptide 2) and retro-inverso (Ac-arg-ser-Gly-ile-tyr-NH2, peptide 3) analogs. Peptide 1 showed the highest mean equilibrated occupancy of the canonical VEGF-A165 pocket (60.3 +/- 34.6%) compared with peptide 2 (25.9 +/- 32.7%) and peptide 3 (21.4 +/- 29.5%), while sampling interactions with canonical pocket residues including Asp320, Tyr297, and Tyr353; FoldX binding-energy estimates were similar across peptides despite their differing pocket persistence. Complementary nanoDSF measurements using recombinant NRP-1 and synthetic peptides yielded apparent dissociation constants in the low micromolar range for all three peptides (12.7 M - 33.3 M), providing experimental support for the predicted interactions. These results support persistent, sequence-dependent association of peptide 1 with the VEGF-A165 binding site of b1-NRP-1 and identify peptide analogs as a candidate for further optimization as an NRP-1-binding peptide.

biophysics↗

bioRxiv2025

Does chlorotoxin target matrix metalloproteinase-2 in glioblastoma?

Glioblastoma aggressively invades surrounding tissue by expressing matrix metalloproteinase-2 (MMP-2). Therefore, effective inhibition of MMP-2 is a desirable target for treatment. In some reports, the chlorotoxin (Ctx) polypeptide produced by the scorpion Leiurus quinquestriatus, interacts with human MMP-2 to inhibit tumor invasion without affecting surrounding tissue. We employed three molecular docking methodologies followed by molecular dynamics simulations to find consensus binding and calculate the binding energy of these peptide ligands to MMP-2. In addition to the Ctx itself, four C-terminal fragments were chosen to study their binding to MMP-2. The molecular docking platforms HPEPDOCK, HADDOCK, and AlphaFold2 created peptide - protein poses for each candidate binding to MMP-2. These poses underwent 500 ns molecular dynamics simulations. Peptide binding on MMP-2 and final binding energies were calculated using the Molecular Mechanics Poisson-Boltzmann Surface Area method. Configurational entropy and root-mean square deviation analyses showed stable peptide - protein complexes. Ctx and its peptide fragments frequently bound to regions on MMP-2 other than the catalytic site. All docking methods shared consensus on large negative binding energies, indicating favorable interaction between Ctx and its analogs with MMP-2. While Ctx and its fragments bind to MMP-2, there is no consensus on which region of MMP-2 they are bound to or which peptide binds strongest. Neither Ctx nor its fragments inhibited MMP-2 enzymatic activity, however, glioblastoma cellular migration was inhibited. Interactions with the non-catalytic regions of MMP-2 suggest allosteric binding to MMP-2. Inhibition of cellular migration without inhibition of MMP-2 activity warrants further study into the possible targets of Ctx expressed in glioblastoma.

biophysics↗

bioRxiv2024

Negative Charges, Not Necessary Phosphorylation, are Required for Ligand Recognition by 14-3-3 Proteins

Protein-protein interactions involving 14-3-3 proteins regulate various cellular activities in normal and pathological conditions. These interactions have mostly been reported to be phosphorylation-dependent, but the 14-3-3 proteins also interact with unphosphorylated proteins. In this work, we investigated whether phosphorylation is required, or, alternatively, whether negative charges are sufficient for 14-3-3{varepsilon} binding. We substituted the pThr residue of pT(502-510) peptide by residues with varying number of negative charges, and investigated binding of the peptides to 14-3-3{varepsilon} using MD simulations and biophysical methods. We demonstrated that at least one negative charge is required for the peptides to bind 14-3-3{varepsilon} while phosphorylation is not necessary, and that two negative charges are preferable for high affinity binding.

biophysics↗

bioRxiv2023

Optimizing Phosphopeptide Structures That Target 14-3-3ϵ in Cutaneous Squamous Cell Carcinoma

14-3-3{varepsilon} is involved in various types of malignancies by increasing cell proliferation, promoting cell invasion or inhibiting apoptosis. In cutaneous squamous cell carcinoma (cSCC), 14-3-3{varepsilon} is over expressed and mislocalized from the nucleus to the cytoplasm where it interacts with the cell division cycle 25 A (CDC25A) and suppresses apoptosis. Hence inhibition of the 14-3-3{varepsilon} - CDC25A interaction is an attractive target for promoting apoptosis in cSCC. In this work, we optimized the structure of our previously designed inhibitor of 14-3-3{varepsilon} - CDC25A interaction, pT, a phosphopeptide fragment corresponding to one of the two binding regions of CDC25A to 14-3-3{varepsilon}. Starting from pT, we developed peptide analogs that bind 14-3-3{varepsilon} with nanomolar affinities. Peptide analogs were designed by shortening the pT peptide, and introducing modifications at position 510 of the pT(502-510) analog. Both molecular dynamics (MD) simulations and biophysical methods were used to determine peptides binding to 14-3-3{varepsilon}. Shortening the pT peptide from 14 to 9 amino acid residues resulted in a peptide (pT(502-510)) that binds 14-3-3{varepsilon} with a KD value of 45.2 nM. Gly to Phe substitution in position 510 of pT(502-510) led to further improvement in affinity (KD: 22.0 nM) of the peptide for 14-3-3{varepsilon}. Our results suggest that the designed peptide analogs are potential candidates for inhibiting 14-3-3{varepsilon} -CDC25A interactions in cSCC cells; thus, inducing their apoptosis.

biochemistry↗

bioRxiv2023

In Silico Transcriptome-based Screens Identify Epidermal Growth Factor Receptor Inhibitors as Therapeutics for Noise-induced Hearing Loss

Noise-Induced Hearing Loss (NIHL) represents a widespread disease for which no therapeutics have been approved by the Food and Drug Administration (FDA). Addressing the conspicuous void of efficacious in vitro or animal models for high throughput pharmacological screening, we utilized an in silico transcriptome-oriented drug screening strategy, unveiling 22 biological pathways and 64 promising small molecule candidates for NIHL protection. Afatinib and zorifertinib, both inhibitors of the Epidermal Growth Factor Receptor (EGFR), were validated for their protective efficacy against NIHL in experimental zebrafish and murine models. This protective effect was further confirmed with EGFR conditional knockout mice and EGF knockdown zebrafish, both demonstrating protection against NIHL. Molecular analysis using Western blot and kinome signaling arrays on adult mouse cochlear lysates unveiled the intricate involvement of several signaling pathways, with particular emphasis on EGFR and its downstream pathways being modulated by noise exposure and Zorifertinib treatment. Administered orally, Zorifertinib was successfully detected in the perilymph fluid of the inner ear in mice with favorable pharmacokinetic attributes. Zorifertinib, in conjunction with AZD5438 - a potent inhibitor of cyclin dependent kinase 2 - produced synergistic protection against NIHL in the zebrafish model. Collectively, our findings underscore the potential application of in silico transcriptome-based drug screening for diseases bereft of efficient screening models and posit EGFR inhibitors as promising therapeutic agents warranting clinical exploration for combatting NIHL. HighlightsO_LIIn silico transcriptome-based drug screens identify pathways and drugs against NIHL. C_LIO_LIEGFR signaling is activated by noise but reduced by zorifertinib in mouse cochleae. C_LIO_LIAfatinib, zorifertinib and EGFR knockout protect against NIHL in mice and zebrafish. C_LIO_LIOrally delivered zorifertinib has inner ear PK and synergizes with a CDK2 inhibitor. C_LI

neuroscience↗

bioRxiv2020

DksA-dependent regulation of RpoS contributes to Borrelia burgdorferi tick-borne transmission and mammalian infectivity

Throughout its enzootic cycle, the Lyme disease spirochete Borreliella (Borrelia) burgdorferi, senses and responds to changes in its environment by using a small repertoire of transcription factors which coordinate the expression of genes required for infection of Ixodes ticks and various mammalian hosts. Among these transcription factors, the DnaK suppressor protein (DksA) plays a pivotal role in regulating gene expression in B. burgdorferi during periods of nutrient limitation and is required for mammalian infectivity. In many pathogenic bacteria, the gene regulatory activity of DksA along with the alarmone guanosine penta- and tetra-phosphate ((p)ppGpp) coordinates the stringent response to various environmental stresses including nutrient limitation. In this study, we sought to characterize the role of DksA in regulating the transcriptional activity of RNA polymerase and in the regulation of RpoS-dependent gene expression required for B. burgdorferi infectivity. Using in vitro transcription assays, we observed recombinant DksA inhibits RpoD-dependent transcription by B. burgdorferi RNA polymerase independent of ppGpp Additionally, we determined the pH-inducible expression of RpoS-dependent genes relies on DksA, but is independent of (p)ppGpp produced by Relbbu. Subsequent transcriptomic and western blot assays indicated DksA regulates the expression of BBD18, a protein previously implicated in the post-transcriptional regulation of RpoS. Moreover, we observed DksA was required for infection of mice following intraperitoneal inoculation or for transmission of B. burgdorferi by Ixodes scapularis nymphs. Together, these data suggest DksA plays a central role in coordinating transcriptional responses of B. burgdorferi required for infectivity through its interactions with RNA polymerase and post-transcriptional control of RpoS. Author SummaryLyme disease, caused by the spirochetal bacteria Borrelia burgdorferi, is the most common vector-borne illness in North America. The ability of B. burgdorferi to establish infection is predicated by its ability to coordinate the expression of virulence factors in response to diverse environmental stimuli encountered within Ixodes ticks and mammalian hosts. Previous studies have shown an essential role for the alternative sigma factor RpoS in regulating the expression of genes required for the successful transmission of B. burgdorferi by Ixodes ticks and infection of mammalian hosts. The DnaK suppressor protein (DksA) is a global gene regulator in B. burgdorferi that also contributes to the expression of RpoS-dependent genes. In this study, we determined DksA exerts its gene regulatory function through direct interactions with the B. burgdorferi RNA polymerase using in vitro transcription assays and controls the expression of RpoS-dependent genes required for mammalian infection by post-transcriptionally regulating cellular levels of RpoS. Our results demonstrate the utility of in vitro transcription assays to determine how gene regulatory proteins like DksA control gene expression in B. burgdorferi, and reveal a novel role for DksA in the infectious cycle of B. burgdorferi.

microbiology↗

bioRxiv2017

Effects of Force Fields on the Conformational and Dynamic Properties of Amyloid β(1-40) Dimer Explored by Replica Exchange Molecular Dynamics Simulations

Alzheimers disease is histologically marked by fibrils of Amyloid beta (A{beta}) peptide within the extracellular matrix. Fibrils themselves are benign compared to the cytotoxicity of the oligomers and pre-fibrillary aggregates. The conformational space and structural ensembles of A{beta} peptides and their oligomers in solution are inherently disordered and proven to be challenging to study. Optimum force field selection for molecular dynamics (MD) simulations and the biophysical relevance of results are still unknown. We compared the conformational space of the A{beta}(1-40) dimers by 300 ns replica exchange MD simulations at physiological temperature (310 K) using: the AMBER-ff99sb-ILDN, AMBER-ff99sb*-ILDN, AMBER-ff99sb-NMR, and CHARMM22* force fields. Statistical comparisons of simulation results to experimental data and previously published simulations utilizing the CHARMM22* and CHARMM36 force fields were performed. All force fields yield sampled ensembles of conformations with collision cross sectional areas for the dimer that are statistically significantly larger than experimental results. All force fields, with the exception of AMBER-ff99sb-ILDN (8.8{+/-}6.4%) and CHARMM36 (2.7{+/-}4.2%), tend to overestimate the -helical content compared to experimental CD (5.3{+/-}5.2%). Using the AMBER-ff99sb-NMR force field resulted in the greatest degree of variance (41.3{+/-}12.9%). Except for the AMBER-ff99sb-NMR force field, the others tended to under estimate the expected amount of {beta}-sheet and over estimate the amount of turn/bend/random coil conformations. All force fields, with the exception AMBER-ff99sb-NMR, reproduce a theoretically expected {beta}-sheet-turn-{beta}-sheet conformational motif, however, only the CHARMM22* and CHARMM36 force fields yield results compatible with collapse of the central and C-terminal hydrophobic cores from residues 17-21 and 30-36. Although analyses of essential subspace sampling showed only minor variations between force fields, secondary structures of lowest energy conformers are different.

biophysics↗