The Entamoeba histolytica,Arp2/3 Complex Is Recruited to Phagocytic Cups through an Atypical Kinase EhAK1

The parasite Entamoeba histolytica is the etiological agent of amoebiasis and phagocytosis plays a key role in virulence of this organism. Signaling pathways involved in activation of cytoskeletal dynamics required for phagocytosis remain to be elucidated. Phagocytosis is initiated with sequential recruitment of EhC2PK, EhCaBP1, EhCaBP3 and an atypical kinase EhAK1 after particle attachment. Here we show that EhARPC1, an essential subunit of the actin branching complex Arp 2/3 is recruited to the phagocytic initiation sites by EhAK1. Imaging, expression knockdown of different molecules and pull down experiments suggest that EhARPC1 interacts with EhAK1 and that it is required during initiation of phagocytosis and phagosome formation. Moreover, recruitment of EhARPC2 at the phagocytosis initiation by EhAK1 is also observed, indicating that the Arp 2/3 complex is recruited. In conclusion, these results suggests a novel mechanism of recruitment of Arp 2/3 complex during phagocytosis in E. histolytica.     

Crystal Structure of a Bivalent Antibody Fab Fragment

We determined the crystal structure to 1.8 Å resolution of the Fab fragment of an affinity-matured human monoclonal antibody (HC84.26.5D) that recognizes the E2 envelope glycoprotein of hepatitis C virus (HCV). Unlike conventional Fabs, which are monovalent monomers, Fab HC84.26.5D assembles into a bivalent domain-swapped dimer in which the two V_L/V_H modules are separated by ~25 Å. In solution, Fab HC84.26.5D exists predominantly as a dimer (~80%) in equilibrium with the monomeric form of the Fab (~20%). Dimerization is mediated entirely by deletion of a single residue, V_HSer113 (Kabat numbering), in the elbow region linking the V_H and C_H1 domains. In agreement with the crystal structure, dimeric Fab HC84.26.5D is able to bind two HCV E2 molecules in solution. This is only the second example of a domain-swapped Fab dimer from among >3000 Fab crystal structures determined to date. Moreover, the architecture of the doughnut-shaped Fab HC84.26.5D dimer is completely different from that of the previously reported Fab 2G12 dimer. We demonstrate that the highly identifiable shape of dimeric Fab HC84.26.5D makes it useful as a fiducial marker for single-particle cryoEM analysis of HCV E2. Bivalent domain-swapped Fab dimers engineered on the basis of HC84.26.5D may also serve as a means of doubling the effective size of conventional Fab–protein complexes for cryoEM.     

Interaction of meropenem with ‘N’ and ‘B’ isoforms of human serum albumin: a spectroscopic and molecular docking study

Carbapenems are used to control the outbreak of β-lactamases expressing bacteria. The effectiveness of drugs is influenced by its interaction with human serum albumin (HSA). Strong binding of carbapenems to HSA may lead to decreased bioavailability of the drug. The non-optimal drug dosage will provide a positive selection pressure on bacteria to develop resistance. Here, we investigated the interaction between meropenem and HSA at physiological pH 7.5 (N-isoform HSA) and non-physiological pH 9.2 (B-isoform HSA). Results showed that meropenem quenches the fluorescence of both ‘N’ and ‘B’ isoforms of HSA (ΔG < 0 and binding constant ~10⁴ M^?1). Electrostatic interactions and van der Waal interactions along with H-bonds stabilized the complex of meropenem with ‘N’ and ‘B’ isoforms of HSA, respectively. Molecular docking results revealed that meropenem binds to HSA near Sudlow’s site II (subdomain IIIA) close to Trp-214 with a contribution of a few residues of subdomain IIA. CD spectroscopy showed a change in the conformation of both the isoforms of HSA upon meropenem binding. The catalytic efficiency of HSA (only N-isoform) on p-nitrophenyl acetate was increased primarily due to a decrease in K_m and an increase in k_cat values. This study provides an insight into the molecular basis of interaction between meropenem and HSA.     

Molecular epidemiology of antibiotic-resistant genes and potent inhibitors against TEM,CTX-M-14, CTX-M-15, and SHV-1 proteins of Escherichia coli in district Peshawar,Pakistan

The Extended Spectrum Beta-Lactamases (ESBLs) producing bacteria is an issue of concern for clinicians resulting in minimize the treatment options. To overcome resistance mechanisms, novel inhibitors with good Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) properties must inhibit the ESBLs resistant genes. The current study aimed to identify the antibiotic resistance genes of ESBLs producing E. coli and a single inhibitor was designed to inhibit all the resistant proteins. The results showed that 42.9% ESBL producers had CTX-M (69.9%), TEM (63.4%), SHV (34.5%) and CTX-M-14 (17.5%) genes. The ESBLs producing isolates were resistant to cephalosporins, quinolones, and sulfonamide with Minimum Inhibitory Concentration (MICs) ranging from 64 to >256 μg/ml. To design multi inhibitory ligands, RECAP synthesis was used for the de-novo discovery of 1000 inhibitors database. Protein crystal structures were retrieved from Protein Data Base (PDB). Lipinski’s rules of five were applied to the novel inhibitors database to improve the ADMET properties. The novel inhibitors database was selected for docking simulations. Placement of the ligand was used by the London dG algorithm implemented in Molecular Operating Environment (MOE), while GBVI/WSA dG algorithm was used for final refinement. Based on docking score, visual inspection of ligands interaction with key residues, binding affinity, and binding energy of ligands with proteins, ten compounds were selected for ESBLs proteins with best ADMET properties, binding energy, and binding affinity the reported ones. These hits compounds have unique scaffolds and are predicted to be a starting point for developing potent inhibitors against antibiotic-resistant proteins.     

Whole-genome sequencing as a first-tier diagnostic framework for rare genetic diseases

Rare diseases affect nearly 300 million people globally with most patients aged five or less. Traditional diagnostic approaches have provided much of the diagnosis; however, there are limitations. For instance, simply inadequate and untimely diagnosis adversely affects both the patient and their families. This review advocates the use of whole genome sequencing in clinical settings for diagnosis of rare genetic diseases by showcasing five case studies. These examples specifically describe the utilization of whole genome sequencing, which helped in providing relief to patients via correct diagnosis followed by use of precision medicine.     

Green synthesis of tellurium-doped SnO2 nanoparticles with sulfurized g-C3N4: Insights into methylene blue photodegradation and antibacterial capability

The construction of SnO₂ nanoparticles (NPs), specifically Te-doped SnO₂ NPs, using a simple and economical co-precipitation technique has been thoroughly described in this work. NH₃ served as the reducing agent in this procedure, whilst polyethylene glycol served as the capping agent. The primary goals of our work were to investigate the physicochemical properties of the synthesized SnO₂ NPs and assess their potential use as antibacterial agents and photocatalysts. Scanning electron microscopy–energy dispersive X-ray, ultraviolet light, Fourier transform infrared spectroscopy, X-ray diffraction (XRD), and other analytical techniques were used to thoroughly analyze the NPs. Based on the full width at half maximum of the most noticeable peaks in the XRD spectrum, the Debye–Scherrer equation was used to calculate the crystallite sizes, which indicated the presence of a single tetragonal SnO₂ phase. Particularly noteworthy was the exceptional photocatalytic activity of graphene-assisted Te-doped SnO₂ NPs, achieving an impressive decomposition efficiency of up to 98% in the photo-oxidation of methylene blue. Furthermore, our investigation delved into the antibacterial attributes of the synthesized SnO₂ NPs against Escherichia coli and Staphylococcus aureus, demonstrating inhibitory effects on both bacteria strains. This suggests potential applications for these NPs in various environmental and medical contexts.     

Novel yellowish-green single-phased spinel Mg1−xBaxAl2O4:Mn2+ phosphor(s) for color rendering white-light-emitting diodes

For white light-rendering research activities, interpretation by using colored emitting materials is an alternative approach. But there are issues in designing the white color emitting materials. Particularly, differences in thermal and decay properties of discrete red, green, and blue emitting materials led to the quest for the search of a single-phased material, able to emit primary colors for white light generation. The current study is an effort to design a simple, single-phase, and cost-effective material with the tunable emission of primary colors by a series of Mg_1−xBa_xAl₂O₄:Mn²⁺ nanopowders. Doping of manganese ion (Mn²⁺) in the presence of the larger barium cation (Ba²⁺) at tetrahedral-sites of the spinel magnesium aluminate (MgAl₂O₄) structure led to the creation of antisite defects. Doped samples were found to have lower bandgaps compared with MgAl₂O₄, and hybridization of 3d-orbitals of Mn²⁺ with O(2p), Mg(2s)/Al(2s3p) was found to be responsible for narrowing the bandgap. The distribution of cations at various sites at random results in a variety of electronic transitions between the valance band and oxygen vacancies as well as electron traps produced the antisite defects. The suggested compositions might be used in white light applications since they have three emission bands with centers at 516 nm (green), 464 nm (blue) and 622 nm (red) at an excitation wavelength of 380 nm. A detailed discussion to analyze the effects of the larger cationic radius of Ba²⁺ on the lattice strain, unit cell parameters, and cell volumes using X-ray diffraction analysis is presented.