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ELife Jul 2024Bacterial exonuclease III (ExoIII), widely acknowledged for specifically targeting double-stranded DNA (dsDNA), has been documented as a DNA repair-associated nuclease...
Bacterial exonuclease III (ExoIII), widely acknowledged for specifically targeting double-stranded DNA (dsDNA), has been documented as a DNA repair-associated nuclease with apurinic/apyrimidinic (AP)-endonuclease and 3'→5' exonuclease activities. Due to these enzymatic properties, ExoIII has been broadly applied in molecular biosensors. Here, we demonstrate that ExoIII () possesses highly active enzymatic activities on ssDNA. By using a range of ssDNA fluorescence-quenching reporters and fluorophore-labeled probes coupled with mass spectrometry analysis, we found ExoIII cleaved the ssDNA at 5'-bond of phosphodiester from 3' to 5' end by both exonuclease and endonuclease activities. Additional point mutation analysis identified the critical residues for the ssDNase action of ExoIII and suggested the activity shared the same active center with the dsDNA-targeted activities of ExoIII. Notably, ExoIII could also digest the dsDNA structures containing 3'-end ssDNA. Considering most ExoIII-assisted molecular biosensors require the involvement of single-stranded DNA (ssDNA) or nucleic acid aptamer containing ssDNA, the activity will lead to low efficiency or false positive outcome. Our study revealed the multi-enzymatic activity and the underlying molecular mechanism of ExoIII on ssDNA, illuminating novel insights for understanding its biological roles in DNA repair and the rational design of ExoIII-ssDNA involved diagnostics.
PubMed: 38959062
DOI: 10.7554/eLife.95648 -
Journal of Chemical Information and... Jul 2024Libraries of collision cross-section (CCS) values have the potential to facilitate compound identification in metabolomics. Although computational methods provide an...
Libraries of collision cross-section (CCS) values have the potential to facilitate compound identification in metabolomics. Although computational methods provide an opportunity to increase library size rapidly, accurate prediction of CCS values remains challenging due to the structural diversity of small molecules. Here, we developed a machine learning (ML) model that integrates graph attention networks and multimodal molecular representations to predict CCS values on the basis of chemical class. Our approach, referred to as MGAT-CCS, had superior performance in comparison to other ML models in CCS prediction. MGAT-CCS achieved a median relative error of 0.47%/1.14% (positive/negative mode) and 1.40%/1.63% (positive/negative mode) for lipids and metabolites, respectively. When MGAT-CCS was applied to real-world metabolomics data, it reduced the number of false metabolite candidates by roughly 25% across multiple sample types ranging from plasma and urine to cells. To facilitate its application, we developed a user-friendly stand-alone web server for MGAT-CCS that is freely available at https://mgat-ccs-web.onrender.com. This work represents a step forward in predicting CCS values and can potentially facilitate the identification of small molecules when using ion mobility spectrometry coupled with mass spectrometry.
PubMed: 38959055
DOI: 10.1021/acs.jcim.3c01934 -
Drugs in R&D Jul 2024Pegfilgrastim-cbqv/CHS-1701 (UDENYCA) (hereafter referred to as pegfilgrastim-cbqv) was approved in 2018 by the US Food and Drug Administration as a biosimilar for...
BACKGROUND
Pegfilgrastim-cbqv/CHS-1701 (UDENYCA) (hereafter referred to as pegfilgrastim-cbqv) was approved in 2018 by the US Food and Drug Administration as a biosimilar for pegfilgrastim (Neulasta) (hereafter referred to as pegfilgrastim). Both pegfilgrastim-cbqv and pegfilgrastim are conjugates of recombinant human granulocyte colony stimulating factor (r-metHuG-CSF) with a 20 kDa polyethylene glycol (PEG) indicated to decrease the incidence of infection, as manifested by febrile neutropenia, in patients receiving myelosuppressive anticancer drugs. The demonstration of analytical similarity for PEG-protein conjugates presents unique challenges since both the protein and PEG attributes must be characterized.
OBJECTIVE
The current study demonstrates the analytical similarity of pegfilgrastim-cbqv and the reference product, pegfilgrastim. In addition to the physicochemical and functional characterization of the protein, the study assessed attributes specific to PEGylation including PEG size and polydispersity, site of attachment, linker composition, and PEGylation process-related variants.
METHODS
The structural, functional, and stability attributes of pegfilgrastim-cbqv and pegfilgrastim were compared using state-of-the-art analytical methods. For the protein, the primary structure, disulfide structure, and secondary and tertiary structures were assessed using traditional protein characterization techniques such as mass spectrometry (MS), circular dichroism (CD), intrinsic fluorescence, and differential scanning calorimetry (DSC), as well as more advanced techniques such as two-dimensional (2D) nuclear magnetic resonance (NMR) and hydrogen deuterium exchange (HDX). For the PEG moiety, the site of attachment, occupancy, linker composition, size and polydispersity were compared using mass spectrometry (both intact and after endoprotease digestion), multiangle light scattering detection (MALS), and Edman degradation. Purity assessments included the assessment of both protein variants and PEGylation variants using chromatographic and electrophoretic analytical separation techniques. The functional similarity between pegfilgrastim-cbqv and pegfilgrastim was compared using both a cell-based bioassay and surface plasmon resonance (SPR). The degradation rates and stability profiles were compared under accelerated and stressed conditions.
RESULTS
Biosimilarity was demonstrated by a thorough assessment of physiochemical and functional attributes, as well as comparative stability, of pegfilgrastim-cbqv relative to pegfilgrastim. These studies demonstrated identical primary structure and disulfide structure, highly similar secondary and tertiary structure, as well as functional similarity. The impurity profile of pegfilgrastim-cbqv was comparable to that of pegfilgrastim with only minor differences in PEGylation variants and a slight offset in the PEG molar mass. These differences were not clinically relevant. The degradation profiles were qualitatively and quantitatively similar under accelerated and stress conditions.
CONCLUSION
The structural, functional, and stability data demonstrate that pegfilgrastim-cbqv is highly similar to the reference product, pegfilgrastim.
PubMed: 38958918
DOI: 10.1007/s40268-024-00471-9 -
Journal of Fluorescence Jul 2024Pedalium Murex leaf extract was used in this study to create Nickel-doped Cerium oxide (Ni-CeO) nanoparticles at 3 mol% and 5 mol% molar concentrations. The...
Pedalium Murex leaf extract was used in this study to create Nickel-doped Cerium oxide (Ni-CeO) nanoparticles at 3 mol% and 5 mol% molar concentrations. The biosynthesized process was applied for the fabrication of Ni-CeO NPs. The X-ray diffraction method was used to identify their crystal structure. The XRD measurements showed that the Ni-CeO NPs crystallized into the face-centred cubic system. Fourier transform infrared spectral study was applied to explore the molecular vibrations and chemical bonding. The surface texture and chemical ingredients of Ni-CeO NPs were studied using field-emission scanning electron microscopy and energy-dispersive X-ray analysis. The EDX mapping spectra illustrate the uniform dispersal of Ce, Ni, and O atoms over the sample's surface. X-ray photoelectron spectroscopy (XPS) was conducted to confirm the chemical state of the Ni-CeO NPs. UV-Vis spectrum study was performed to ascertain the photon absorption, bandgap, and Urbach edge of Ni-CeO NPs. Photoluminescence (PL) research has been used to study the light-emitting characteristic of Ni-CeO NPs. The emissive intensity transition corresponding to Ni-CeO NPs was found to increase with the dopant level. The CIE 1931 chromaticity map was plotted to find the aptness of the samples for optical uses. The antifungal ability of Ni-CeO NPs was evaluated against the fungi candida albicans and candida krusein with the agar well-diffusion process. The fungicidal activity of the 3 mol% Ni doped CeO nanoparticles has shown a maximum zone of inhibition. The experimental findings illustrate the utility of Ni-CeO NPs for optical and antifungal applications.
PubMed: 38958905
DOI: 10.1007/s10895-024-03831-5 -
European Journal of Clinical... Jul 2024Leminorella grimontii strain LG-KP-E1-2-T0 was isolated from Zophobas morio larvae. It showed a susceptibility phenotype compatible with the expression of an inducible...
Leminorella grimontii strain LG-KP-E1-2-T0 was isolated from Zophobas morio larvae. It showed a susceptibility phenotype compatible with the expression of an inducible extended-spectrum β-lactamase. The presence of a chromosomal bla gene encoding for the class A GRI-1 β-lactamase was revealed by whole-genome sequencing. GRI-1 shared the highest amino acid identity with RIC-1 and OXY-type β-lactamases (76-80%). Analysis of six further publicly-available L. grimontii draft genomes deposited in NCBI revealed that bla was always present. Core-genome analysis indicated that LG-KP-E1-2-T0 was unique from other strains. We provided the first complete genome of L. grimontii and new insights on its chromosomal β-lactamases.
PubMed: 38958811
DOI: 10.1007/s10096-024-04888-7 -
Journal of Neurology Jul 2024Aquaporin-4 (AQP4) antibody-associated neuromyelitis optica spectrum disorder (NMOSD) is an antibody-mediated inflammatory disease of the central nervous system. We have... (Review)
Review
BACKGROUND
Aquaporin-4 (AQP4) antibody-associated neuromyelitis optica spectrum disorder (NMOSD) is an antibody-mediated inflammatory disease of the central nervous system. We have undertaken a systematic review and meta-analysis to ascertain the sex ratio and mean age of onset for AQP4 antibody associated NMOSD. We have also explored factors that impact on these demographic data.
METHODS
A systematic search of databases was conducted according to the PRISMA guidelines. Articles reporting sex distribution and age of onset for AQP4 antibody-associated NMSOD were reviewed. An initially inclusive approach involving exploration with regression meta-analysis was followed by an analysis of just AQP4 antibody positive cases.
RESULTS
A total of 528 articles were screened to yield 89 articles covering 19,415 individuals from 88 population samples. The female:male sex ratio was significantly influenced by the proportion of AQP4 antibody positive cases in the samples studied (p < 0.001). For AQP4 antibody-positive cases the overall estimate of the sex ratio was 8.89 (95% CI 7.78-10.15). For paediatric populations the estimate was 5.68 (95% CI 4.01-8.03) and for late-onset cases, it was 5.48 (95% CI 4.10-7.33). The mean age of onset was significantly associated with the mean life expectancy of the population sampled (p < 0.001). The mean age of onset for AQP4 antibody-positive cases in long-lived populations was 41.7 years versus 33.3 years in the remainder.
CONCLUSIONS
The female:male sex ratio and the mean age of onset of AQP4 antibody-associated NMOSD are significantly higher than MS. The sex ratio increases with the proportion of cases that are positive for AQP4 antibodies and the mean age of onset increases with population life expectancy.
PubMed: 38958756
DOI: 10.1007/s00415-024-12452-8 -
Applied Microbiology and Biotechnology Jul 2024This study investigated the potential of endophytic fungi to produce paclitaxel (Taxol®), a potent anticancer compound widely employed in chemotherapy. This research...
This study investigated the potential of endophytic fungi to produce paclitaxel (Taxol®), a potent anticancer compound widely employed in chemotherapy. This research aimed to identify, confirm, and characterize endophytic fungi capable of paclitaxel (PTX) production and assess their paclitaxel yield. Additionally, it aimed to investigate factors influencing paclitaxel production. A total of 100 endophytic fungal isolates were collected and identified from the roots of Artemisia judaica. Aspergillus fumigatiaffinis exhibited the highest PTX production (26.373 μg L) among the isolated endophytic fungi. The strain was identified as A. fumigatiaffinis (Accession No. PP235788.1). Molecular identification confirmed its novelty, representing the first report of PTX production by A. fumigatiaffinis, an endophyte of Artemisia judaica. Optimization through full factorial design of experiments (DOE) and response surface methodology (RSM) significantly enhanced PTX production to 110.23 μg L from 1 g of dry weight of the fungal culture under optimal conditions of pH 8.0, 150 μg L becozyme supplementation, and 18 days of fermentation in potato dextrose broth. The presence of paclitaxel was confirmed using thin layer chromatography, high performance liquid chromatography, and gas chromatography-mass spectrometry. These findings maximize the role of endophytic fungus to produce a secondary metabolite that might be able to replace the chemically produced PTX and gives an opportunity to provide a sustainable source of PTX eco-friendly at high concentrations. KEY POINTS: • Endophytic fungi, like A. fumigatiaffinis, show promise for eco-friendly paclitaxel production • Optimization strategies boost paclitaxel yield significantly, reaching 110.23 μg L • Molecular identification confirms novelty, offering a sustainable PTX source.
Topics: Paclitaxel; Aspergillus; Endophytes; Fermentation; Plant Roots; Culture Media; Gas Chromatography-Mass Spectrometry; Chromatography, High Pressure Liquid
PubMed: 38958755
DOI: 10.1007/s00253-024-13230-2 -
Organic Letters Jul 2024Antiaromatic nucleophilic substitution reactions in cycloheptatrienide pyridinium and phosphonium zwitterions with initial formation of a cycloheptatetraene intermediate...
Antiaromatic nucleophilic substitution reactions in cycloheptatrienide pyridinium and phosphonium zwitterions with initial formation of a cycloheptatetraene intermediate are explored. The mechanism was supported by quantum chemical calculations, first-order reaction kinetics, and high-resolution mass spectrometry. The pyridinium zwitterion exhibited weak antiaromaticity, whereas the intermediate displayed Möbius aromaticity, as evidenced by nuclear independent chemical shift values and the shape of its HOMO. This study represents the eighth confirmed instance of a Möbius-aromatic organic species in its ground state.
PubMed: 38958743
DOI: 10.1021/acs.orglett.4c01446 -
A Feeding Trial to Investigate Strategies to Mitigate the Impacts of Poisoning in Australian Cattle.Journal of Agricultural and Food... Jul 2024poisoning of cattle causes distinct symptoms and frequently death, attributable to the toxin simplexin. poisoning was induced via addition of ground plant to the...
poisoning of cattle causes distinct symptoms and frequently death, attributable to the toxin simplexin. poisoning was induced via addition of ground plant to the daily feed in a three-month trial with Droughtmaster steers. The trial tested four potential mitigation treatments, namely, biochar, activated biochar, bentonite, and a bacterial inoculum, and incorporated negative and positive control groups. All treatments tested were unable to prevent the development of simplexin poisoning effects. However, steers consuming a bentonite adsorbent together with showed lesser rates-of-decline for body weight ( < 0.05) and four hematological parameters ( < 0.02), compared to the positive control group fed only. Microbiome analysis revealed that despite displaying poisoning symptoms, the rumen microbial populations of animals receiving were very resilient, with dominant bacterial populations maintained over time. Unexpectedly, clinical edema developed in some animals up to 2 weeks after dosing was ceased.
PubMed: 38958707
DOI: 10.1021/acs.jafc.4c02082 -
Advanced Science (Weinheim,... Jul 2024Increasing evidence suggests the role of reactive oxygen and nitrogen species (RONS) in regulating antitumor immune effects and immunosuppression. RONS modify...
Increasing evidence suggests the role of reactive oxygen and nitrogen species (RONS) in regulating antitumor immune effects and immunosuppression. RONS modify biomolecules and induce oxidative post-translational modifications (oxPTM) on proteins that can alarm phagocytes. However, it is unclear if and how protein oxidation by technical means could be a strategy to foster antitumor immunity and therapy. To this end, cold gas plasma technology producing various RONS simultaneously to oxidize the two melanoma-associated antigens MART and PMEL is utilized. Cold plasma-oxidized MART (oxMART) and PMEL (oxPMEL) are heavily decorated with oxPTMs as determined by mass spectrometry. Immunization with oxidized MART or PMEL vaccines prior to challenge with viable melanoma cells correlated with significant changes in cytokine secretion and altered T-cell differentiation of tumor-infiltrated leukocytes (TILs). oxMART promoted the activity of cytotoxic central memory T-cells, while oxPMEL led to increased proliferation of cytotoxic effector T-cells. Similar T-cell results are observed after incubating splenocytes of tumor-bearing mice with B16F10 melanoma cells. This study, for the first time, provides evidence of the importance of oxidative modifications of two melanoma-associated antigens in eliciting anticancer immunity.
PubMed: 38958560
DOI: 10.1002/advs.202404131