Exploring the Impact of C-Terminal Based Pentapeptides on the Disassembly of Aß42 Fibrils.

An effective therapeutic strategy to suppress Alzheimer's disease (AD) progression is to disrupt ß-sheet rich neurotoxic soluble amyloid-ß (Aß) aggregates. Previously, we identified new pentapeptides (RVVPI and RIAPA) with notably enhanced ability to block Aß42 aggregation as compared to Aß42 C-terminal derived peptide RIIGL using integrated computational protocol. In this work, the potential of RIIGL, RVVPI, and RIAPA for the structural destabilization of Aß42 protofibril was assessed by molecular dynamics (MD) simulations and in vitro studies. The binding free energy analysis depicts that charged residues influence Aß42 protofibril-pentapeptide interactions. Notably, RVVPI displays a more pronounced destabilization effect than other peptides due to higher conformational fluctuations, and disruption of salt bridge (K28-A42) interactions in Aß42 protofibril. RVVPI exhibited highest inhibitory activity (Inhibition= 66.2%, IC50= 5.57 ± 0.83 µM) against Aß42 aggregation consistent with computational results. Remarkably, RVVPI displayed ~4.5 fold lower IC50 value as compared to RIIGL. ThT and TEM studies highlighted the enhanced efficiency of RVVPI (62.4%) in the disassembly of pre-formed Aß42 fibrils than RIIGL and RIAPA. The combined in silico and in vitro studies identified a new peptide, RVVPI, as an efficient inhibitor of Aß42 fibrillation and disassembly of Aß42 aggregates.

Nano-Biotechnology for Bacteria Identification and Potent Anti-bacterial Properties: A Review of Current State of the Art.

Sepsis is a critical disease caused by the abrupt increase of bacteria in human blood, which subsequently causes a cytokine storm. Early identification of bacteria is critical to treating a patient with proper antibiotics to avoid sepsis. However, conventional culture-based identification takes a long time. Polymerase chain reaction (PCR) is not so successful because of the complexity and similarity in the genome sequence of some bacterial species, making it difficult to design primers and thus less suitable for rapid bacterial identification. To address these issues, several new technologies have been developed. Recent advances in nanotechnology have shown great potential for fast and accurate bacterial identification. The most promising strategy in nanotechnology involves the use of nanoparticles, which has led to the advancement of highly specific and sensitive biosensors capable of detecting and identifying bacteria even at low concentrations in very little time. The primary drawback of conventional antibiotics is the potential for antimicrobial resistance, which can lead to the development of superbacteria, making them difficult to treat. The incorporation of diverse nanomaterials and designs of nanomaterials has been utilized to kill bacteria efficiently. Nanomaterials with distinct physicochemical properties, such as optical and magnetic properties, including plasmonic and magnetic nanoparticles, have been extensively studied for their potential to efficiently kill bacteria. In this review, we are emphasizing the recent advances in nano-biotechnologies for bacterial identification and anti-bacterial properties. The basic principles of new technologies, as well as their future challenges, have been discussed.

Multiepitope glycan based laser assisted fluorescent nanocomposite with dual functionality for sensing and ablation of Pseudomonas aeruginosa.

Pseudomonas aeruginosa (P. aeruginosa) infection is becoming a severe health hazard and needs early diagnosis with high specificity. However, the non-specific binding of a biosensor is a challenge to the current bacterial detection system. For the first time, we chemically synthesized a galactose tripod (GT) as a P. aeruginosa-specific ligand. We conjugated GT to a photothermally active fluorescent nanocomposite (Au@SiO2-TCPP). P. aeruginosa can be detected using Au@SiO2-TCPP-GT, and additionally ablated as well using synergistic photothermal and photodynamic therapy. Molecular dynamics and simulation studies suggested better binding of GT (binding energy = -6.6 kcal mol-1) with P. aeruginosa lectin than that of galactose monopod (GM) (binding energy = -5.9 kcal mol-1). Furthermore, a binding study was extended to target P. aeruginosa, which has a galactose-binding carbohydrate recognition domain receptor. The colorimetric assay confirmed a limit of detection (LOD) of 104 CFU mL-1. We also looked into the photosensitizing property of Au@SiO2-TCPP-GT, which is stimulated by laser light (630 nm) and causes photoablation of bacteria by the formation of singlet oxygen in the surrounding media. The cytocompatibility of Au@SiO2-TCPP-GT was confirmed using cytotoxicity assays on mammalian cell lines. Moreover, Au@SiO2-TCPP-GT also showed non-hemolytic activity. Considering the toxicity analysis and efficacy of the synthesized glycan nanocomposites, these can be utilized for the treatment of P. aeruginosa-infected wounds. Furthermore, the current glycan nanocomposites can be used for bacterial detection and ablation of P. aeruginosa in contaminated food and water samples as well.

Salmonella typhimurium detection and ablation using OmpD specific aptamer with non-magnetic and magnetic graphene oxide.

Foodborne diseases have increased in the last few years due to the increased consumption of packaged and contaminated food. Major foodborne bacteria cause diseases such as diarrhea, vomiting, and sometimes death. So, there is a need for early detection of foodborne bacteria as pre-existing detection techniques are time-taking and tedious. Aptamer has gained interest due to its high stability, specificity, and sensitivity. Here, aptamer has been developed against Salmonella Typhimurium through the Cell-Selex method, and to further find the reason for specificity and sensitivity, OmpD protein was isolated, and binding studies were done. Single molecular FRET experiment using aptamer and graphene oxide studies has also been done to understand the mechanism of FRET and subsequently used for target bacterial detection. Using this assay, Salmonella Typhimurium can be detected up to 10 CFU/mL. Further, Magnetic Graphene oxide was used to develop an assay to separate and ablate bacteria using 808 nm NIR where temperature increase was more than 60 °C within 30 s and has been shown by plating as well as a confocal live dead assay. Thus, using various techniques, bacteria can be detected and ablated using specific aptamer and Graphene oxide.

Triazole-Peptide Conjugate as a Modulator of Aß-Aggregation, Metal-Mediated Aß-Aggregation, and Cytotoxicity.

Amyloid-ß (Aß) aggregation plays a key role in the pathogenesis of Alzheimer's disease (AD). Along with this, the presence of redox-active metals like Cu2+ further enhances Aß aggregation, oxidative stress, and cellular toxicity. In this study, we have rationally designed, synthesized, and evaluated a series of triazole-peptide conjugates as potential promiscuous ligands capable of targeting different pathological factors of AD. In particular, peptidomimetic DS2 showed the best inhibitory activity against Aß aggregation with an IC50 value of 2.43 ± 0.05 µM. In addition, DS2 disaggregates preformed Aß42 fibrils, chelates metal ions, inhibits metal-mediated Aß aggregation, significantly controls reactive oxygen species production, and reduces oxidative stress. DS2 exhibited very low cytotoxicity and significantly ameliorated the Aß-induced toxicity in differentiated neuroblastoma cells, SH-SY5Y. In addition, alteration in the fibrillary architecture of Aß42 in the absence and presence of DS2 was validated by transmission electron microscopy (TEM) images. To shed light on the inhibitory mechanism of DS2 against Aß aggregation and disassembly of the protofibril structure, molecular dynamics (MD) simulations have been performed. DS2 binds preferentially with the central hydrophobic core (CHC) residues of Aß42 monomer and chains D-E of Aß42 protofibril. The dictionary of secondary structure of proteins analysis indicated a noteworthy increase in the helix content from 38.5 to 61% and, notably, a complete loss of ß-sheet content of Aß42 monomer when DS2 is added to it. DS2 suppressed Aß42 monomer aggregation by preserving helical conformations and was able to reduce the production of aggregation-prone ß-sheet structures, which are consistent with ThT, circular dichroism, and TEM assay that indicate a reduction in the formation of toxic Aß42 aggregated species on the addition of DS2. Moreover, DS2 destabilized the Aß42 protofibril structure by significantly reducing the binding affinity between chains D-E of protofibril, which highlighted the disruption of interchain interactions and subsequent deformation of the protofibril structure. The results of the present study demonstrate that triazole-peptide conjugates may be valuable chemotypes for the development of promising multifunctional AD therapeutic candidates.

Mechanobactericidal, Gold Nanostar Hydrogel-Based Bandage for Bacteria-Infected Skin Wound Healing.

The emergence of multidrug resistant (MDR) microorganisms has led to the development of alternative approaches for providing relief from microbial attacks. The mechano-bactericidal action as a substitute for antimicrobials has become the focus of intensive research. In this work, nanostructure-conjugated hydrogel are explored as a flexible dressing against Staphylococcus aureus (S. aureus)-infected skin wounds. Herein gold nanostars (AuNst) with spike lengths reaching 120 nm are probed for antibacterial action. The bacterial killing of >95% is observed for Pseudomonas aeruginosa (P. aeruginosa) and Escherichia coli (E. coli), while up to 60% for Gram-positive S. aureus. AuNst conjugated hydrogel (AuNst120@H) reduced >80% colonies of P. aeruginosa and E. coli. In comparison, around 35.4% reduction of colonies are obtained for S. aureus. The viability assay confirmed the presence of about 85% of living NIH-3T3 cells when grown with hydrogels. An animal wound model is also developed to assess the efficiency of AuNst120@H. A significant reduction in wound size is observed on the 10th day in AuNst120@H treated animals with fully formed epidermal layers, hair follicles, new blood vessels, and arrector muscles. These findings suggest that novel dressing materials can be developed with antimicrobial nanotextured surfaces.

Nanoglycocluster based diagnostic platform for colorimetric detection of bacteria; A comparative study analysing the effect of AuNPs size, linker length, and glycan diversity.

Nanoglycoclusters, an upcoming class of functional nanomaterial are known to drive various processes like detection, imaging, targeting proteins, cells, and bacteria. Nanoglycoclusters are a type of nanomaterial functionalized with various glycans. The array of glycan in multiple copies enhances binding affinity with proteins. Selective and sensitive bacteria/lectin interactions using nanomaterials are an emerging area of research. The measurement of different ligand receptor interactions require sophisticated analytical tools that limit the application in biosensor domain. Recently, colorimetric biosensors gained importance in the field of the biosensor for the detection of bacteria/lectins. Herein we have demonstrated that different size of gold nanoparticles (AuNPs) along with various polyethylene glycol (PEG) linkers, functionalized with synthesized monopod and tripod of mannose and galactose that have different bacteria/lectins specificity. The newly synthesized nanoglycoclusters were able to discriminate between different lectins and bacteria. The aggregation of specific nanoglycocluster upon interaction with specific bacteria/lectins revealed that mannose monopod (MM) and mannose tripod (MT) are specific to Escherichia coli and concanavalin A (ConA) lectin, while galactose monopod (GM) and galactose tripod (GT) are specific to Pseudomonas aeruginosa and Peanut agglutinin (PNA) lectin. Further, the binding events depict the affinity of tripod glycans is more with respect to its corresponding monopod glycans. Our findings explored the potential of colorimetric sensing depending upon the size of AuNPs, linker length, specificity, along with glycans density to develop user friendly diagnostic system for the detection of bacteria.

1,2,3-Triazole ß-lactam conjugates as antimicrobial agents.

A convenient and efficient synthesis of new triazole ß-lactam conjugates using click chemistry is described. ß-lactam 15 and 16 were prepared using cycloaddition strategy and propargylated at N-1 to afford compounds 17 and 18. Cu-catalyzed click reaction of these ß-lactams 17 and 18 with different aryl azides provided 1,2,3-triazole conjugates 6 and 7, respectively. The products were fully characterized spectroscopically and tested against Gram-(+) and Gram-(-) bacteria. Compound 7a and 7c were found to be most active.

Multifunctional Mono-Triazole Derivatives Inhibit Aß42 Aggregation and Cu2+-Mediated Aß42 Aggregation and Protect Against Aß42-Induced Cytotoxicity.

Amyloid beta (Aß) peptide aggregation is considered as one of the key hallmarks of Alzheimer's disease (AD). Moreover, Aß peptide aggregation increases considerably in the presence of metal ions and triggers the generation of reactive oxygen species (ROS), which ultimately leads to oxidative stress and neuronal damage. Based on the 'multitarget-directed ligands' (MTDLs) strategy, we designed, synthesized, and evaluated a novel series of triazole-based compounds for AD treatment via experimental and computational methods. Among the designed MTDLs [4(a-x)], the triazole derivative 4v exhibited the most potent inhibition of self-induced Aß42 aggregation (78.02%) with an IC50 value of 4.578 ± 0.109 µM and also disassembled the preformed Aß42 aggregates significantly. In addition, compound 4v showed excellent metal chelating ability and maintained copper in the redox-dormant state to prevent the generation of ROS in copper-ascorbate redox cycling. Further, 4v significantly inhibited Cu2+-induced Aß42 aggregation and disassembled the Cu2+-induced Aß42 protofibrils as compared to the reference compound clioquinol (CQ). Importantly, 4v did not show cytotoxicity and was able to inhibit the toxicity induced by Aß42 aggregates in SH-SY5Y cells. Molecular docking results confirmed the strong binding of 4v with Aß42 monomer and Aß42 protofibril structure. The experimental and molecular docking results highlighted that 4v is a promising multifunctional lead compound for AD.

Multi-target-directed triazole derivatives as promising agents for the treatment of Alzheimer's disease.

A novel series of triazole-based compounds have been designed, synthesised and evaluated as multi-target-directed ligands (MTDLs) against Alzheimer disease (AD). The triazole-based compounds have been designed to target four major AD hallmarks that include Aß aggregation, metal-induced Aß aggregation, metal dys-homeostasis and oxidative stress. Among the synthesised compounds, 6n having o-CF3 group on the phenyl ring displayed most potent inhibitory activity (96.89% inhibition, IC50 = 8.065 ±â€¯0.129 µM) against Aß42 aggregation, compared to the reference compound curcumin (95.14% inhibition, IC50 = 6.385 ±â€¯0.009 µM). Compound 6n disassembled preformed Aß42 aggregates as effectively as curcumin. Furthermore, 6n displayed metal chelating ability and significantly inhibited Cu2+-induced Aß42 aggregation and disassembled preformed Cu2+-induced Aß42 aggregates. 6n successfully controlled the generation of the reactive oxygen species (ROS) by preventing the copper redox cycle. In addition, 6n did not display cytotoxicity and was able to inhibit toxicity induced by Aß42 aggregates in SH-SY5Y cells. The preferred binding regions and key interactions of 6n with Aß42 monomer and Aß42 protofibril structure was evaluated with molecular docking. Compound 6n binds preferably to the C-terminal region of Aß42 that play a critical role in Aß42 aggregation. The results of the present study highlight a novel triazole-based compound, 6n, as a promising MTDL against AD.

Role of novel multidrug efflux pump involved in drug resistance in Klebsiella pneumoniae.

BACKGROUND: Multidrug resistant Klebsiella pneumoniae have caused major therapeutic problems worldwide due to the emergence of the extended-spectrum ß-lactamase producing strains. Although there are >10 major facilitator super family (MFS) efflux pumps annotated in the genome sequence of the K. pneumoniae bacillus, apparently less is known about their physiological relevance. PRINCIPAL FINDINGS: Insertional inactivation of kpnGH resulting in increased susceptibility to antibiotics such as azithromycin, ceftazidime, ciprofloxacin, ertapenem, erythromycin, gentamicin, imipenem, ticarcillin, norfloxacin, polymyxin-B, piperacillin, spectinomycin, tobramycin and streptomycin, including dyes and detergents such as ethidium bromide, acriflavine, deoxycholate, sodium dodecyl sulphate, and disinfectants benzalkonium chloride, chlorhexidine and triclosan signifies the wide substrate specificity of the transporter in K. pneumoniae. Growth inactivation and direct fluorimetric efflux assays provide evidence that kpnGH mediates antimicrobial resistance by active extrusion in K. pneumoniae. The kpnGH isogenic mutant displayed decreased tolerance to cell envelope stressors emphasizing its added role in K. pneumoniae physiology. CONCLUSIONS AND SIGNIFICANCE: The MFS efflux pump KpnGH involves in crucial physiological functions besides being an intrinsic resistance determinant in K. pneumoniae.