Interaction between Yersinia pestis Ail Outer Membrane Protein and the C-Terminal Domain of Human Vitronectin.

Yersinia pestis, the causative agent of plague, is capable of evading the human immune system response by recruiting the plasma circulating vitronectin proteins, which act as a shield and avoid its lysis. Vitronectin recruitment is mediated by its interaction with the bacterial transmembrane protein Ail, protruding from the Y. pestis outer membrane. By using all-atom long-scale molecular dynamic simulations of Ail embedded in a realistic model of the bacterial membrane, we have shown that vitronectin forms a stable complex, mediated by interactions between the disordered moieties of the two proteins. The main amino acids driving the complexation have also been evidenced, thus favoring the possible rational design of specific peptides which, by inhibiting vitronectin recruitment, could act as original antibacterial agents.

Synthesis and preliminary anticancer evaluation of photo-responsive prodrugs of hydroxymethylene bisphosphonate alendronate.

The antitumoral activity of hydroxymethylene bisphosphonates (HMBP) such as alendronate or zoledronate is hampered by their exceptional bone-binding properties and their short plasmatic half-life which preclude their accumulation in non-skeletal tumors. In this context, the use of lipophilic prodrugs represents a simple and straightforward strategy to enhance the biodistribution of bisphosphonates in these tissues. We describe in this article the synthesis of light-responsive prodrugs of HMBP alendronate. These prodrugs include lipophilic photo-removable nitroveratryl groups which partially mask the highly polar alendronate HMBP scaffold. Photo-responsive prodrugs of alendronate are stable in physiological conditions and display reduced toxicity compared to alendronate against MDA-MB-231 cancer cells. However, the antiproliferative effect of these prodrugs is efficiently restored after cleavage of their nitroveratryl groups upon exposure to UV light. In addition, substitution of alendronate with such photo-responsive substituents drastically reduces its bone-binding properties, thereby potentially improving its biodistribution in soft tissues after i.v. administration. The development of such lipophilic photo-responsive prodrugs is a promising approach to fully exploit the anticancer effect of HMBPs on non-skeletal tumors.

The antibacterial properties of branched peptides based on poly(l-arginine): In vitro antibacterial evaluation and molecular dynamic simulations.

The emergence of bacterial strains resistant to antibiotics is a major issue in the medical field. Antimicrobial peptides are widely studied as they do not generate as much resistant bacterial strains as conventional antibiotics and present a broad range of activity. Among them, the homopolypeptide poly(l-arginine) presents promising antibacterial properties, especially in the perspective of its use in biomaterials. Linear poly(l-arginine) has been extensively studied but the impact of its 3D structure remains unknown. In this study, the antibacterial properties of newly synthesized branched poly(l-arginine) peptides, belonging to the family of multiple antigenic peptides, are evaluated. First, in vitro activities of the peptides shows that branched poly(l-arginine) is more efficient than linear poly(l-arginine) containing the same number of arginine residues. Surprisingly, peptides with more arms and more residues are not the most effective. To better understand these unexpected results, interactions between these peptides and the membranes of Gram positive and Gram negative bacteria are simulated thanks to molecular dynamic. It is observed that the bacterial membrane is more distorted by the branched structure than by the linear one and by peptides containing smaller arms. This mechanism of action is in full agreement with in vitro results and suggest that our simulations form a robust model to evaluate peptide efficiency towards pathogenic bacteria.

DockSurf: A Molecular Modeling Software for the Prediction of Protein/Surface Adhesion.

The elucidation of structural interfaces between proteins and inorganic surfaces is a crucial aspect of bionanotechnology development. Despite its significance, the interfacial structures between proteins and metallic surfaces are yet to be fully understood, and the lack of experimental investigation has impeded the development of many devices. To overcome this limitation, we suggest considering the generation of protein/surface structures as a molecular docking problem with a homogenous plan as the target. To this extent, we propose a new software, DockSurf, which aims to quickly propose reliable protein/surface structures. Our approach considers the conformational exploration with Euler's angles, which provide a cartography instead of a unique structure. Interaction energies were derived from quantum mechanics computations for a set of small molecules that describe protein atom types and implemented in a Derjaguin, Landau, Verwey, and Overbeek potential for the consideration of large systems such as proteins. The validation of DockSurf software was conducted with molecular dynamics for corona proteins with gold surfaces and provided enthusiastic results. This software is implemented in the RPBS platform to facilitate widespread access to the scientific community.

Mechanism of the Covalent Inhibition of Human Transmembrane Protease Serine 2 as an Original Antiviral Strategy.

The Transmembrane Protease Serine 2 (TMPRSS2) is a human enzyme which is involved in the maturation and post-translation of different proteins. In addition to being overexpressed in cancer cells, TMPRSS2 plays a further fundamental role in favoring viral infections by allowing the fusion of the virus envelope with the cellular membrane, notably in SARS-CoV-2. In this contribution, we resort to multiscale molecular modeling to unravel the structural and dynamical features of TMPRSS2 and its interaction with a model lipid bilayer. Furthermore, we shed light on the mechanism of action of a potential inhibitor (nafamostat), determining the free-energy profile associated with the inhibition reaction and showing the facile poisoning of the enzyme. Our study, while providing the first atomistically resolved mechanism of TMPRSS2 inhibition, is also fundamental in furnishing a solid framework for further rational design targeting transmembrane proteases in a host-directed antiviral strategy.

Theobroma cacao improves bone growth by modulating defective ciliogenesis in a mouse model of achondroplasia.

A gain-of-function mutation in the fibroblast growth factor receptor 3 gene (FGFR3) results in achondroplasia (ACH), the most frequent form of dwarfism. Constitutive activation of FGFR3 impairs bone formation and elongation and many signal transduction pathways. Identification of new and relevant compounds targeting the FGFR3 signaling pathway is of broad importance for the treatment of ACH, and natural plant compounds are prime drug candidate sources. Here, we found that the phenolic compound (-)-epicatechin, isolated from Theobroma cacao, effectively inhibited FGFR3's downstream signaling pathways. Transcriptomic analysis in an Fgfr3 mouse model showed that ciliary mRNA expression was modified and influenced significantly by the Indian hedgehog and PKA pathways. (-)-Epicatechin is able to rescue mRNA expression impairments that control both the structural organization of the primary cilium and ciliogenesis-related genes. In femurs isolated from a mouse model (Fgfr3Y367C/+) of ACH, we showed that (-)-epicatechin eliminated bone growth impairment during 6 days of ex vivo culture. In vivo, we confirmed that daily subcutaneous injections of (-)-epicatechin to Fgfr3Y367C/+ mice increased bone elongation and rescued the primary cilium defects observed in chondrocytes. This modification to the primary cilia promoted the typical columnar arrangement of flat proliferative chondrocytes and thus enhanced bone elongation. The results of the present proof-of-principle study support (-)-epicatechin as a potential drug for the treatment of ACH.

Between Two Walls: Modeling the Adsorption Behavior of ß-Glucosidase A on Bare and SAM-Functionalized Gold Surfaces.

The efficient immobilization of enzymes on surfaces remains a complex but central issue in the biomaterials field, which requires us to understand this process at the atomic level. Using a multiscale approach combining all-atom molecular dynamics and coarse-grain Brownian dynamics simulations, we investigated the adsorption behavior of ß-glucosidase A (ßGA) on bare and self-assembled monolayer (SAM)-functionalized gold surfaces. We monitored the enzyme position and orientation during the molecular dynamics (MD) trajectories and measured the contacts it forms with both surfaces. While the adsorption process has little impact on the protein conformation, it can nonetheless perturb its mechanical properties and catalytic activity. Our results show that compared to the SAM-functionalized surface, the adsorption of ßGA on bare gold is more stable, but less specific, and more likely to disrupt the enzyme's function. This observation emphasizes the fact that the structural organization of proteins at the solid interface is a key point when designing devices based on enzyme immobilization, as one must find an acceptable stability-activity trade-off.

Computational Studies of a DNA-Based Aptasensor: toward Theory-Driven Transduction Improvement.

Aptamers are a class of bioreceptors intensively used in current analytical tools dedicated to molecular diagnostics due to their ability to perform large structural reorganization upon target binding. However, there is a lack of methodologies allowing us to rationalize their structure in order to improve the transduction efficiency in aptamer sensors. We choose here, as a model system, a three-strand DNA structure as the probe, composed of two DNA strands anchored on a gold surface and partially hybridized with an aptamer sequence sensitive to ampicillin (AMP). The DNA structure has been designed to show strong structural change upon AMP binding to its aptamer. Using a set of computational techniques including molecular dynamics simulations, we deeply investigated the structure change upon analyte binding, taking into account the grafting on the surface. Original analyses of ion distributions along the trajectories unveil a distinct pattern between both states which can be related to changes in capacitance of the interface between these states. To our knowledge, this work demonstrates the ability of computational investigations for the first time to drive, in silico, the design of aptasensors.

Grafting TRAIL through Either Amino or Carboxylic Groups onto Maghemite Nanoparticles: Influence on Pro-Apoptotic Efficiency.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF cytokine superfamily. TRAIL is able to induce apoptosis through engagement of its death receptors DR4 and DR5 in a wide variety of tumor cells while sparing vital normal cells. This makes it a promising agent for cancer therapy. Here, we present two different ways of covalently grafting TRAIL onto maghemite nanoparticles (NPs): (a) by using carboxylic acid groups of the protein to graft it onto maghemite NPs previously functionalized with amino groups, and (b) by using the amino functions of the protein to graft it onto NPs functionalized with carboxylic acid groups. The two resulting nanovectors, NH-TRAIL@NPs-CO and CO-TRAIL@NPs-NH, were thoroughly characterized. Biological studies performed on human breast and lung carcinoma cells (MDA-MB-231 and H1703 cell lines) established these nanovectors are potential agents for cancer therapy. The pro-apoptotic effect is somewhat greater for CO-TRAIL@NPs-NH than NH-TRAIL@NPs-CO, as evidenced by viability studies and apoptosis analysis. A computational study indicated that regardless of whether TRAIL is attached to NPs through an acid or an amino group, DR4 recognition is not affected in either case.

Synthesis and preliminary anticancer evaluation of new triazole bisphosphonate-based isoprenoid biosynthesis inhibitors.

The synthesis of a new set of triazole bisphosphonates 8a-d and 9a-d presenting an alkyl or phenyl substituent at the C-4 or C-5 position of the triazole ring is described. These compounds have been evaluated for their antiproliferative activity against MIA PaCa-2 (pancreas), MDA-MB-231 (breast) and A549 (lung) human tumor cell lines. 4-hexyl- and 4-octyltriazole bisphosphonates 8b-c both displayed remarkable antiproliferative activities with IC50 values in the micromolar range (0.75-2.4 µM) and were approximately 4 to 12-fold more potent than zoledronate. Moreover, compound 8b inhibits geranylgeranyl pyrophosphate biosynthesis in MIA PaCa-2 cells which ultimately led to tumor cells death.

First dual binder of microRNA-146a and monomeric tau: a novel approach for multitargeted therapeutics for neurodegenerative diseases.

We report a new approach for the development of multitargeted therapeutics for Alzheimer's disease (AD) based on dual targeting of monomeric tau and biogenesis of microRNA-146a. Compound MG-1102 displayed a superior neuroprotective activity, in comparison to mono-targeted therapeutics, which validates the likelihood of the success of this approach in AD drug development.

Computational approach and electrochemical measurements for protein detection with MIP-based sensor.

Rapid and accurate detection of proteins in biological fluids is increasingly required in the biomedical environment. Actually, it is performed with conventional techniques, which are generally run by robotized platforms at centralized laboratories. In this work, molecular dynamics calculations and an experimental procedure were conducted to set up electrochemical sensors based on polypyrrol (PPy) molecular imprinted polymers (MIP) for proteins detection. Here, prostate-specific antigen (PSA) was selected as a template model. Computational calculations indicate that for any PPy conformation and any amino-acid location in the protein, PSA molecules remain strongly inserted in the PPy polymer without biological alterations. One from possible orientations, appeared to be most probable as it presents the lowest absorption energy (-363 kcal mol-1) and largest contact area (4034.1 Å2). The device was then elaborated by in situ electropolymerization of PPy films. MIP's thickness and extraction duration were optimized by chronoamperometry. Square wave voltammetry technique was investigated for PSA detection in standard solution in the concentration range of 3x10 -8 ng.ml-1- 300 ng ml-1. According to the Hill equation, the equilibrium dissociation constant Kdbetween PSA and its imprint was estimated at Kd = (1.02 ±â€¯0.54) × 10-14 M, confirming the strong binding between the designed MIP and the protein as predicted by the computational study. PSA concentration values directly measured in 35 human serum samples were found closely correlated to those measured by the ELISA technique. The promising fast and low-cost sensor might be used successfully for proteins detection at low concentrations with high selectivity and reproducibility.

In Vitro interaction between yeast frataxin and superoxide dismutases: Influence of mitochondrial metals.

BACKGROUND: Friedreich's ataxia results from a decreased expression of the nuclear gene encoding the mitochondrial protein, frataxin. Frataxin participates in the biosynthesis of iron-sulfur clusters and heme cofactors, as well as in iron storage and protection against oxidative stress. How frataxin interacts with the antioxidant defence components is poorly understood. METHODS: Therefore, we have investigated by kinetic, thermodynamic and modelling approaches the molecular interactions between yeast frataxin (Yfh1) and superoxide dismutases, Sod1 and Sod2, and the influence of Yfh1 on their enzymatic activities. RESULTS: Yfh1 interacts with cytosolic Sod1 with a dissociation constant, Kd = 1.3 ±â€¯0.3 µM, in two kinetic steps. The first step occurs in the 200 ms range and corresponds to the Yfh1-Sod1 interaction, whereas the second is slow and is assumed to be a change in the conformation of the protein-protein adduct. Furthermore, computational investigations confirm the stability of the Yfh1-Sod1 complex. Yfh1 forms two protein complexes with mitochondrial Sod2 with 1:1 and 2:1 Yfh1/Sod2 stoichiometry (Kd1 = 1.05 ±â€¯0.05 and Kd2 = 6.6 ±â€¯0.1 µM). Furthermore, Yfh1 increases the enzymatic activity of Sod1 while slightly affecting that of Sod2. Finally, the stabilities of the protein-protein adducts and the effect of Yfh1 on superoxide dismutase activities depend on the nature of the mitochondrial metal. CONCLUSIONS: This work confirms the participation of Yfh1 in cellular defence against oxidative stress.

Molecular Dynamics Simulation of a RNA Aptasensor.

Single-stranded RNA aptamers have emerged as novel biosensor tools. However, the immobilization procedure of the aptamer onto a surface generally induces a loss of affinity. To understand this molecular process, we conducted a complete simulation study for the Flavin mononucleotide aptamer for which experimental data are available. Several molecular dynamics simulations (MD) of the Flavin in complex with its RNA aptamer were conducted in solution, linked with six thymidines (T6) and, finally, immobilized on an hexanol-thiol-functionalized gold surface. First, we demonstrated that our MD computations were able to reproduce the experimental solution structure and to provide a meaningful estimation of the Flavin free energy of binding. We also demonstrated that the T6 linkage, by itself, does not generate a perturbation of the Flavin recognition process. From the simulation of the complete biosensor system, we observed that the aptamer stays oriented parallel to the surface at a distance around 36 Å avoiding, this way, interaction with the surface. We evidenced a structural reorganization of the Flavin aptamer binding mode related to the loss of affinity and induced by an anisotropic distribution of sodium cationic densities. This means that ionic diffusion is different between the surface and the aptamer than above this last one. We suggest that these findings might be extrapolated to other nucleic acids systems for the future design of biosensors with higher efficiency and selectivity.

Tyrosine kinase inhibitor NVP-BGJ398 functionally improves FGFR3-related dwarfism in mouse model.

Achondroplasia (ACH) is the most frequent form of dwarfism and is caused by gain-of-function mutations in the fibroblast growth factor receptor 3-encoding (FGFR3-encoding) gene. Although potential therapeutic strategies for ACH, which aim to reduce excessive FGFR3 activation, have emerged over many years, the use of tyrosine kinase inhibitor (TKI) to counteract FGFR3 hyperactivity has yet to be evaluated. Here, we have reported that the pan-FGFR TKI, NVP-BGJ398, reduces FGFR3 phosphorylation and corrects the abnormal femoral growth plate and calvaria in organ cultures from embryos of the Fgfr3Y367C/+ mouse model of ACH. Moreover, we demonstrated that a low dose of NVP-BGJ398, injected subcutaneously, was able to penetrate into the growth plate of Fgfr3Y367C/+ mice and modify its organization. Improvements to the axial and appendicular skeletons were noticeable after 10 days of treatment and were more extensive after 15 days of treatment that started from postnatal day 1. Low-dose NVP-BGJ398 treatment reduced intervertebral disc defects of lumbar vertebrae, loss of synchondroses, and foramen-magnum shape anomalies. NVP-BGJ398 inhibited FGFR3 downstream signaling pathways, including MAPK, SOX9, STAT1, and PLCγ, in the growth plates of Fgfr3Y367C/+ mice and in cultured chondrocyte models of ACH. Together, our data demonstrate that NVP-BGJ398 corrects pathological hallmarks of ACH and support TKIs as a potential therapeutic approach for ACH.

Striking HIV-1 Entry by Targeting HIV-1 gp41. But, Where Should We Target?

HIV-1 gp41 facilitates the viral fusion through a conformational switch involving the association of three C-terminal helices along the conserved hydrophobic grooves of three N-terminal helices coiled-coil. The control of these structural rearrangements is thought to be central to HIV-1 entry and, therefore, different strategies of intervention are being developed. Herewith, we describe a procedure to simulate the folding of an HIV-1 gp41 simplified model. This procedure is based on the construction of plausible conformational pathways, which describe protein transition between non-fusogenic and fusogenic conformations. The calculation of the paths started with 100 molecular dynamics simulations of the non-fusogenic conformation, which were found to converge to different intermediate states. Those presenting defined criteria were selected for separate targeted molecular dynamics simulations, subjected to a force constant imposing a movement towards the gp41 fusogenic conformation. Despite significant diversity, a preferred sequence of events emerged when the simulations were analyzed in terms of the formation, breakage and evolution of the contacts. We pointed out 29 residues as the most relevant for the movement of gp41; also, 2696 possible interactions were reduced to only 48 major interactions, which reveals the efficiency of the method. The analysis of the evolution of the main interactions lead to the detection of four main behaviors for those contacts: stable, increasing, decreasing and repulsive interactions. Altogether, these results suggest a specific small cavity of the HIV-1 gp41 hydrophobic groove as the preferred target to small molecules.

A novel small-molecule inhibitor of HIV-1 entry.

BACKGROUND: Antiretroviral therapy has transformed HIV-1 infection into a managed condition with near-normal life expectancy. However, a significant number of patients remain with limited therapeutic options due to HIV-1 resistance, side effects, or drug costs. Further, it is likely that current drugs will not retain efficacy, due to risks of side effects and transmitted resistance. RESULTS: We describe compound 5660386 (3-ethyl-2-[3-(1,3,3-trimethyl-1,3-dihydro-2H-indol-2-ylidene)-1-propen-1-yl]-1,3-benzothiazol-3-ium) as a novel inhibitor of HIV-1 entry. Compound 5660386 inhibits HIV-1 entry in cell lines and primary cells, binds to HIV-1 envelope protein, and inhibits the interaction of GP120 to CD4. Further, compound 5660386 showed a unique and broad-range activity against primary HIV-1 isolates from different subtypes and geographical areas. CONCLUSION: Development of small-molecule entry inhibitors of HIV-1 such as 5660386 may lead to novel classes of anti-HIV-1 therapeutics. These inhibitors may be particularly effective against viruses resistant to current antiretroviral drugs and could have potential applications in both treatment and prevention.

Simulation with quantum mechanics/molecular mechanics for drug discovery.

INTRODUCTION: Biological macromolecules, such as proteins or nucleic acids, are (still) molecules and thus they follow the same chemical rules that any simple molecule follows, even if their size generally renders accurate studies unhelpful. However, in the context of drug discovery, a detailed analysis of ligand association is required for understanding or predicting their interactions and hybrid quantum mechanics/molecular mechanics (QM/MM) computations are relevant tools to help elucidate this process. AREAS COVERED: In this review, the authors explore the use of QM/MM for drug discovery. After a brief description of the molecular mechanics (MM) technique, the authors describe the subtractive and additive techniques for QM/MM computations. The authors then present several application cases in topics involved in drug discovery. EXPERT OPINION: QM/MM have been widely employed during the last decades to study chemical processes such as enzyme-inhibitor interactions. However, despite the enthusiasm around this area, plain MM simulations may be more meaningful than QM/MM. To obtain reliable results, the authors suggest fixing several keystone parameters according to the underlying chemistry of each studied system.

Molecular modeling study of the induced-fit effect on kinase inhibition: the case of fibroblast growth factor receptor 3 (FGFR3).

Tyrosine kinases are a wide family of targets with strong pharmacological relevance. These proteins undergo large-scale conformational motions able to inactivate them. By the end of one of these structural processes, a new cavity is opened allowing the access to a specific type of inhibitors, called type II. The kinase domain of fibroblast growth factor receptor 3 (FGFR3) falls into this family of kinases. We describe here, for the first time, its inactivation process through target molecular dynamics. The transient cavity, at the crossroad between the DFGout and Cα helix out inactivation is herein explored. Molecular docking calculations of known ligands demonstrated that type II inhibitors are able to interact with this metastable transient conformation of FGFR3 kinase. Besides, supplemental computations were conducted and clearly show that type II inhibitors drive the kinase inactivation process through specific stabilization with the DFG triad. This induced-fit effect of type II ligands toward FGFR3 might be extrapolated to other kinase systems and provides meaningful structural information for future drug developments.

Indole alkaloids from the Marquesan plant Rauvolfia nukuhivensis and their effects on ion channels.

In addition to the already reported nukuhivensiums 1 and 2, 11 indole alkaloids were isolated from the bark of the plant Rauvolfia nukuhivensis, growing in the Marquesas archipelago. The known sandwicine (3), isosandwicine (4), spegatrine (8), lochneram (9), flavopereirine (13) have been found in this plant together with the norsandwicine (5), isonorsandwicine (6), Nb-methylisosandwicine (7), 10-methoxypanarine (10), nortueiaoine (11), tueiaoine (12). The structure elucidation was performed on the basis of a deep exploration of the NMR and HRESIMS data as well as comparison with literature data for similar compounds. Norsandwicine, 10-methoxypanarine, tueiaoine, and more importantly nukuhivensiums, were shown to significantly induce a reduction of IKr amplitude (HERG current). Molecular modelling through docking was performed in order to illustrate this result.