Early stages of misfolding of PAP248-286 at two different pH values: An insight from molecular dynamics simulations

PAP248-286 peptides, which are highly abundant in human semen, aggregate and form amyloid fibrils that enhance HIV infection. Previous experimental studies have shown that the infection-promoting activity of PAP248-286 begins to increase well before amyloid formation takes place and that pH plays a key role in the enhancement of PAP248-286-related infection. Hence, understanding the early stages of misfolding of the PAP2482-86 peptide is crucial. To this end, we have performed 60 independent MD simulations for a total of 24 µs at two different pH values (4.2 and 7.2). Our data shows that early stages of misfolding of the PAP248-286 peptide is a multistage process and that the first step of the process is a transition from an "I-shaped" structure to a "U-shaped" structure. We further observed that the structure of PAP248-286 at the two different pH values shows significantly different features. At pH 4.2, the peptide has less intra-molecular H-bonds and a reduced α-helical content than at pH 7.2. Moreover, differences in intra-peptide residues contacts are also observed at the two pH values. Finally, free energy landscape analysis shows that there are more local minima in the energy surface of the peptide at pH 7.2 than at pH 4.2. Overall, the present study elucidates the early stages of misfolding of the PAP248-286 peptide at the atomic level, thus possibly opening new avenues in structure-based drug discovery against HIV infection

Crystal Structure of the Apo and the ADP-Bound Form of Choline Kinase from Plasmodium falciparum

Among the malaria-causing parasites, the deadliest isPlasmodium falciparum, which accounts for the majority of the fatalities. As the infection progresses inside erythrocytes, major cellular and metabolic changes take place. For its own growth, the parasite relies on the accumulation of phospholipids, which are essential for membrane synthesis. Within the Kennedy pathway, theP. falciparumcholine kinase (PfChoK) has a central role in the biosynthesis of phosphatidylcholine and its selective inhibition leads to the parasite arrest and eradication. Here, we report the crystal structure of the apo and the ADP-bound form of choline kinase fromPlasmodium falciparumat 2.0 and 2.2 angstrom resolution, respectively. These new structural data will facilitate the implementation of effective structure-based drug development strategies against PfChoK in the fight against malaria

Recent Approaches to the Identification of Novel Microtubule-Targeting Agents

Microtubules are key components of the eukaryotic cytoskeleton with essential roles in cell division, intercellular transport, cell morphology, motility, and signal transduction. They are composed of protofilaments of heterodimers of alpha-tubulin and beta-tubulin organized as rigid hollow cylinders that can assemble into large and dynamic intracellular structures. Consistent with their involvement in core cellular processes, affecting microtubule assembly results in cytotoxicity and cell death. For these reasons, microtubules are among the most important targets for the therapeutic treatment of several diseases, including cancer. The vast literature related to microtubule stabilizers and destabilizers has been reviewed extensively in recent years. Here we summarize recent experimental and computational approaches for the identification of novel tubulin modulators and delivery strategies. These include orphan small molecules, PROTACs as well as light-sensitive compounds that can be activated with high spatio-temporal accuracy and that represent promising tools for precision-targeted chemotherapy

Hybrid 1D Plasmonic/Photonic Crystals are Responsive to Escherichia Coli

Photonic crystal-based biosensors hold great promise as valid and low-cost devices for real-time monitoring of a variety of biotargets. Given the high processability and easiness of read-out even for unskilled operators, these systems can be highly appealing for the detection of bacterial contaminants in food and water. Here, we propose a novel hybrid plasmonic/photonic device that is responsive to Escherichia coli, which is one of the most hazardous pathogenic bacterium. Our system consists of a thin layer of silver, a metal that exhibits both a plasmonic behavior and a well-known biocidal activity, on top of a solution processed 1D photonic crystal. We attribute the bio-responsivity to the modification of the dielectric properties of the silver film upon bacterial contamination, an effect that likely stems from the formation of polarization charges at the Ag/bacterium interface within a sort of bio-doping mechanism. Interestingly, this triggers a blue-shift in the photonic response. This work demonstrates that our hybrid plasmonic/photonic device can be a low-cost and portable platform for the detection of common contaminants in food and water

Biohybrid Electrospun Membrane for the Filtration of Ketoprofen Drug from Water

A current challenge in materials science and biotechnology is to express a specific and controlled functionality on the large interfacial area of a nanostructured material to create smart biohybrid systems for targeted applications. Here, we report on a biohybrid system featuring poly(vinyl alcohol) as the supporting synthetic polymer and bovine serum albumin as the biofunctional element. The optimal processing conditions to produce these self-standing composite membranes are determined, and the composition and distribution of the bioactive agent within the polymeric matrices are analyzed. A post-processing cross-linking using glutaraldehyde enables this functional membrane to be used as a chemical filter in aqueous environments. By demonstrating that our mats can remove large amounts of ketoprofen from water, we show that the combination of a BSA-induced biofunctionality with a nanostructured fibrous material allows for the development of an efficient biohybrid filtering device for the large and widely used family of nonsteroidal anti-inflammatory drugs (NSAIDs). The crystal structure of the complex between BSA and ketoprofen is determined for the first time and confirms the interaction between the two species

Molecular dynamics simulations provide insights into the substrate specificity of FAOX family members

Enzymatic assays based on Fructosyl Amino Acid Oxidases (FAOX) represent a potential, rapid and economical strategy to measure glycated hemoglobin (HbA1c), which is in turn a reliable method to monitor the insurgence and the development of diabetes mellitus. However, the engineering of naturally occurring FAOX to specifically recognize fructosyl-valine (the glycated N-terminal residue of HbA1c) has been hindered by the paucity of information on the tridimensional structures and catalytic residues of the different FAOX that exist in nature, and in general on the molecular mechanisms that regulate specificity in this class of enzymes. In this study, we use molecular dynamics simulations and advanced modeling techniques to investigate five different relevant wild-type FAOX (Amadoriase I, Amadoriase II, PnFPOX, FPOX-E and N1-1-FAOD) in order to elucidate the molecular mechanisms that drive their specificity towards polar and nonpolar substrates. Specifically, we compare these five different FAOX in terms of overall folding, ligand entry tunnels, ligand binding residues and ligand binding energies. Our work will contribute to future enzyme structure modifications aimed at the rational design of novel biosensors for the monitoring of blood glucose levels

Synthesis and evaluation of an agrocin 84 toxic moiety (TM84) analogue as a malarial threonyl tRNA synthetase inhibitor

An analogue of a toxic moiety (TM84) of natural product agrocin 84 containing threonine amide instead of 2,3-dihydroxy-4-methylpentanamide was prepared and evaluated as a putative Plasmodium falciparum threonyl t-RNA synthetase (PfThrRS) inhibitor. This TM84 analogue features submicromolar inhibitory potency (IC50 = 440 nM) comparable to that of borrelidin (IC50 = 43 nM) and therefore complements chemotypes known to inhibit malarial PfThrRS, which are currently limited to borrelidin and its analogues. The crystal structure of the inhibitor in complex with the E. coli homologue enzyme (EcThrRS) was obtained, revealing crucial ligand-protein interactions that will pave the way for the design of novel ThrRS inhibitors

A combined fragment-based virtual screening and STD-NMR approach for the identification of E-cadherin ligands

Cadherins promote cell-cell adhesion by forming homophilic interactions via their N-terminal extracellular domains. Hence, they have broad-ranging physiological effects on tissue organization and homeostasis. When dysregulated, cadherins contribute to different aspects of cancer progression and metastasis; therefore, targeting the cadherin adhesive interface with small-molecule antagonists is expected to have potential therapeutic and diagnostic value. Here, we used molecular docking simulations to evaluate the propensity of three different libraries of commercially available drug-like fragments (nearly 18,000 compounds) to accommodate into the Trp2 binding pocket of E-cadherin, a crucial site for the orchestration of the protein’s dimerization mechanism. Top-ranked fragments featuring five different aromatic chemotypes were expanded by means of a similarity search on the PubChem database (Tanimoto index >90%). Of this set, seven fragments containing an aromatic scaffold linked to an aliphatic chain bearing at least one amine group were finally selected for further analysis. Ligand-based NMR data (Saturation Transfer Difference, STD) and molecular dynamics simulations suggest that these fragments can bind E-cadherin mostly through their aromatic moiety, while their aliphatic portions may also diversely engage with the mobile regions of the binding site. A tetrahydro-β-carboline scaffold functionalized with an ethylamine emerged as the most promising fragment

The interplay of soft-hard substituents in photochromic diarylethenes

A series of diarylethenes with substituents of different size and chemical nature was synthesised showing that beside some intermolecular interactions involving the central diarylethene core, lateral groups clearly play a key role in the crystal packing arrangements. These structural features were further analyzed in relation to the thermal data obtained by differential scanning calorimetry (DSC) and monitored using FT-IR spectroscopy, thus providing a rationalization of the observed thermal transitions processes. The role of van der Waals interactions is crucial in driving crystal packing formation towards loosely packed arrangements characterized by large hydrophobic contact areas. Interestingly, some functional substituents favour an amorphous state after thermal treatment, a peculiar feature that can be exploited to design uniform photochromic layers

Tailoring FPOX enzymes for enhanced stability and expanded substrate recognition

Abstract Fructosyl peptide oxidases (FPOX) are deglycating enzymes that find application as key enzymatic components in diabetes monitoring devices. Indeed, their use with blood samples can provide a measurement of the concentration of glycated hemoglobin and glycated albumin, two well-known diabetes markers. However, the FPOX currently employed in enzymatic assays cannot directly detect whole glycated proteins, making it necessary to perform a preliminary proteolytic treatment of the target protein to generate small glycated peptides that can act as viable substrates for the enzyme. This is a costly and time consuming step. In this work, we used an in silico protein engineering approach to enhance the overall thermal stability of the enzyme and to improve its catalytic activity toward large substrates. The final design shows a marked improvement in thermal stability relative to the wild type enzyme, a distinct widening of its access tunnel and significant enzymatic activity towards a range of glycated substrates