Notes

Fingerprinting vanadium within soils based on speciation characteristics as well as isotope compositions.
4 mM) of MVA was successfully produced in 48 h without any addition of nutrients except methane. Our platform exhibited high stability and reproducibility with regard to cell growth and MVA production. We believe that this versatile system can be easily extended to many other value-added processes and has a variety of potential applications.We have previously reported the unique features of dimeric bisaminoquinolines as anticancer agents and have identified their cellular target as PPT1, a protein palmitoyl-thioesterase. We now report a systematic study on the role of the linker in these constructs, both with respect to the distance between the heterocycles, the linker hydrophobicity and the methylation status (primary vs. secondary vs. tertiary) of the central nitrogen atom on the observed biological activity.The enzyme 2-methylerythritol 2,4-cyclodiphosphate synthase, IspF, is essential for the biosynthesis of isoprenoids in most bacteria, some eukaryotic parasites, and the plastids of plant cells. The development of inhibitors that target IspF may lead to novel classes of anti-infective agents or herbicides. Enantiomers of tryptophan hydroxamate were synthesized and evaluated for binding to Burkholderia pseudomallei (Bp) IspF. The L-isomer possessed the highest potency, binding BpIspF with a KD of 36 µM and inhibited BpIspF activity 55% at 120 µM. The high-resolution crystal structure of the L-tryptophan hydroxamate (3)/BpIspF complex revealed a non-traditional mode of hydroxamate binding where the ligand interacts with the active site zinc ion through the primary amine. In addition, two hydrogen bonds are formed with active site groups, and the indole group is buried within the hydrophobic pocket composed of side chains from the 60 s/70 s loop. Along with the co-crystal structure, STD NMR studies suggest the methylene group and indole ring are potential positions for optimization to enhance binding potency.Machado-Joseph disease (MJD) is a fatal neurodegenerative disorder clinically characterized by prominent ataxia. It is caused by an expansion of a CAG trinucleotide in the ATXN3 gene, translating into an expanded polyglutamine (polyQ) tract in the ATXN3 protein, that becomes prone to misfolding and aggregation. The pathogenesis of the disease has been associated with the dysfunction of several cellular mechanisms, including autophagy and transcription regulation. Here we investigated the transcriptional modifications of the autophagy pathway in models of MJD and assessed whether modulating the levels of the affected autophagy-associated transcripts (AATs), would alleviate MJD-associated pathology. Our results show that autophagy is impaired at the transcriptional level in MJD, affecting multiple AATs, including Unc-51 like autophagy activating kinase 1 and 2 (ULK1 and ULK2), two homologs involved in autophagy induction. Reinstating ULK1/2 levels by AAV-mediated gene transfer, significantly improved motor performance, while preventing neuropathology in two in vivo models of MJD. Moreover, in vitro studies showed that the observed positive effects may be mainly attributed to ULK1 activity. This study provides a strong evidence of the beneficial effect of ULK homologs overexpression, suggesting these as promising instruments for the treatment of MJD and other neurodegenerative disorders.Moderate noise exposure may cause acute loss of cochlear synapses without affecting the cochlear hair cells and hearing threshold; thus, it remains "hidden" to standard clinical tests. This cochlear synaptopathy is one of the main pathologies of noise-induced hearing loss (NIHL). There is no effective treatment for NIHL, mainly because of the lack of a proper drug delivery technique. We hypothesized that local magnetic delivery of gene therapy into the inner ear could be beneficial for NIHL. In this study, we used superparamagnetic iron oxide nanoparticles (SPIONs) and a recombinant adeno associated virus vector (AAV2(quad Y-F)) to deliver brain-derived neurotrophic factor (BDNF)-gene therapy into the rat inner ear via minimally invasive magnetic targeting. We found that the magnetic targeting effectively accumulates and distributes SPION tagged AAV2(quad Y-F)-BDNF vector into the inner ear. We also found that AAV2(quad Y-F) efficiently transfects cochlear hair cells and enhances BDNF gene expression. Enhanced BDNF gene expression substantially recovers noise-induced BDNF gene downregulation, ABR wave I amplitude reduction, and synapse loss. These results suggest that magnetic targeting of AAV2(quad Y-F)-mediated BDNF gene therapy could reverse cochlear synaptopathy after NIHL.Prime editor (PE), a new genome editing tool, can generate all 12 possible base-to-base conversions, insertion, and deletion of short fragment DNA. PE has the potential to correct the majority of known human genetic disease-related mutation. AAVs, the safe vector widely used in clinic, are not capable of delivering PE (∼6.3kb) in a single vector because of the limited loading capacity (∼4.8 kb). To accommodate the loading capacity of AAVs, we constructed four split-PE (split-PE994, split-PE1005, split-PE1024, and split-PE1032) using Rma intein. Using a GFP mutated reporter system, PE reconstituting activities were screened, and two efficient split-PEs (split-PE1005 and split-PE1024) were identified. We then demonstrated that split-PEs delivered by dual-AAV1, especially split-PE1024, could mediate base transversion and insertion at four endogenous sites in human cells. To test the performance of split-PE in vivo, split-PE1024 was then delivered into the adult mouse retina by dual-AAV8. We demonstrated successful editing of Dnmt1 in adult mouse retina. Our study provides a new method to deliver PE to adult tissue, paving the way for in vivo gene editing therapy using PE.Non-human primates (NHPs) are a preferred animal model for optimizing adeno-associated virus (AAV)-mediated CNS gene delivery protocols before clinical trials. In spite of its inherent appeal, it is challenging to compare different serotypes, delivery routes, and disease indications in a well-powered, comprehensive, multigroup NHP experiment. Here, a multiplex barcode recombinant AAV (rAAV) vector-tracing strategy has been applied to a systemic analysis of 29 distinct, wild-type (WT), AAV natural isolates and engineered capsids in the CNS of eight macaques. The report describes distribution of each capsid in 15 areas of the macaques' CNS after intraparenchymal (putamen) injection, or cerebrospinal fluid (CSF)-mediated administration routes (intracisternal, intrathecal, or intracerebroventricular). To trace the vector biodistribution (viral DNA) and targeted tissues transduction (viral mRNA) of each capsid in each of the analyzed CNS areas, quantitative next-generation sequencing analysis, assisted by the digital-droplet PCR technology, was used. The report describes the most efficient AAV capsid variants targeting specific CNS areas after each route of administration using the direct side-by-side comparison of WT AAV isolates and a new generation of rationally designed capsids. The newly developed bioinformatics and visualization algorithms, applicable to the comparative analysis of several mammalian brain models, have been developed and made available in the public domain.Individuals must constantly adjust their behavior to adapt to the changing environment, and this dynamic adjustment ability has been studied through the conflict adaptation effect (CAE). We explored the resting state brain network underlying individual differences in CAE. The functional connection strength between dorsolateral prefrontal cortex (DLPFC) and bilateral insula or right anterior cingulate cortex (ACC) was negatively correlated with CAE, whereas the connection strength between DLPFC and left inferior parietal lobule was positively correlated with CAE. We speculated that, when faced with conflict, individuals with low CAE employ salience network to detect conflict, and then control is performed by DLPFC; in individuals with high CAEs who maintain a high level of control in all tasks, conflict is resolved mainly by the central executive network. In other words, in conflict tasks, individuals with high CAEs adopt proactive control strategies, but their counterparts with low CAEs employ reactive control strategies.Anthrax, by Bacillus anthracis, remains a dreadful fatal hazard worldwide. The currently used anthrax vaccines are plagued by numerous issues that limit their widespread use. As an immunization approach targeting both extracellular antigens and toxins of B. anthracis may achieve better sterile immunity, the present investigation designed a bicistronic secretory anti-anthrax DNA vaccine targeting immune response against toxin and cells. The efficacy of the vaccine was compared with monocistronic DNA vaccines and the currently used anthrax vaccine. For this, mice were immunized with the developed vaccine containing pag (encoding protective antigen to block toxin) and eag genes (encoding EA1 to target cells) of B. anthracis through DNA-prime/Protein-boost (D/P) and DNA prime/DNA-boost (D/D) approaches. There was a >2 and > 5 fold increase in specific antibody level by D/D and D/P approaches respectively, on 42nd days post-immunization (dpi). Serum cytokine profiling showed that both Th1 and Th2 immune responses were elicited, with more Th2 responses in D/P strategy. More importantly, challenge with 100 times LD50 of B. anthracis at 42nd dpi exhibited maximum cumulative survival (83.33 %) by bicistronic D/P approach. HDAC inhibitor Remarkably, immunization with EA1 delayed mortality onset in infection. The study forms the first report on complement-dependent bactericidal activity of antiEA1 antibodies. In short, co-immunization of PA and EA1 through the developed bicistronic DNA vaccine would be an effective immunization approach in anthrax vaccination. Further, D/P strategy could enhance vaccine-induced immunity against B. anthracis. Altogether, the study generates certain critical insights having direct applications in next-generation vaccine development against anthrax.With the occurrence and evolution of antibiotic and multidrug resistance in bacteria most of the existing remedies are becoming ineffective. The pan-proteome exploration of the bacterial pathogens helps to identify the wide spectrum therapeutic targets which will be effective against all strains in a species. The current study is focused on the pan-proteome profiling of zoonotic pathogen Orientia tsutsugamushi (Ott) for the identification of potential therapeutic targets. The pan-proteome of Ott is estimated to be extensive in nature that has 1429 protein clusters, out of which 694 were core, 391 were accessory, and 344 were unique. It was revealed that 622 proteins were essential, 222 proteins were virulent factors, and 42 proteins were involved in antibiotic resistance. The potential therapeutic targets were further classified into eleven broad classes among which gene expression and regulation, transport, and metabolism were dominant. The biological interactome analysis of therapeutic targets revealed that an ample amount of interactions were present among the proteins involved in DNA replication, ribosome assembly, cellwall metabolism, cell division, and antimicrobial resistance. The predicted therapeutic targets from the pan-proteome of Ott are involved in various biological processes, virulence, and antibiotic resistance; hence envisioned as potential candidates for drug discovery to combat scrub typhus.
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