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PeerJ 2023The rapid emergence of antibiotic-resistant bacteria directly contributes to a wave of untreatable infections. The lack of new drug development is an important driver of...
BACKGROUND
The rapid emergence of antibiotic-resistant bacteria directly contributes to a wave of untreatable infections. The lack of new drug development is an important driver of this crisis. Most antibiotics today are small molecules that block vital processes in bacteria. To optimize such effects, the three-dimensional structure of targeted bacterial proteins is imperative, although such a task is time-consuming and tedious, impeding the development of antibiotics. The development of RNA-based therapeutics has catalyzed a new platform of antibiotics-antisense oligonucleotides (ASOs). These molecules hybridize with their target mRNAs with high specificity, knocking down or interfering with protein translation. This study aims to develop a bioinformatics pipeline to identify potent ASO targets in essential bacterial genes.
METHODS
Three bacterial species (, , and ) were used to demonstrate the utility of the pipeline. Open reading frames of bacterial essential genes were downloaded from the Database of Essential Genes (DEG). After filtering for specificity and accessibility, ASO candidates were ranked based on their self-hybridization score, predicted melting temperature, and the position on the gene in an operon. Enrichment analysis was conducted on genes associated with putative potent ASOs.
RESULTS
A total of 45,628 ASOs were generated from 348 unique essential genes in . A total of 1,117 of them were considered putative. A total of 27,273 ASOs were generated from 191 unique essential genes in . A total of 847 of them were considered putative. A total of 175,606 ASOs were generated from 346 essential genes in . A total of 7,061 of them were considered putative. Critical biological processes associated with these genes include translation, regulation of cell shape, cell division, and peptidoglycan biosynthetic process. Putative ASO targets generated for each bacterial species are publicly available here: https://github.com/EricSHo/AOA. The results demonstrate that our bioinformatics pipeline is useful in identifying unique and accessible ASO targets in bacterial species that post major public health issues.
Topics: Oligonucleotides, Antisense; Anti-Bacterial Agents; Staphylococcus aureus; Oligonucleotides
PubMed: 38025700
DOI: 10.7717/peerj.16343 -
Biosensors Dec 2023Immunoassays based on antibodies as recognizing elements and enzymes as signal-generating modules are extensively used now in clinical lab diagnostics, food, and... (Review)
Review
Immunoassays based on antibodies as recognizing elements and enzymes as signal-generating modules are extensively used now in clinical lab diagnostics, food, and environmental analyses. However, the application of natural enzymes and antibodies has some drawbacks, such as relatively high manufacturing costs, thermal instability, and lot-to-lot variations that lower the reproducibility of results. Oligonucleotide aptamers are able to specifically bind their targets with high affinity and selectivity, so they represent a prospective alternative to protein antibodies for analyte recognition. Their main advantages include thermal stability and long shelf life, cost-efficient chemical synthesis, and negligible batch-to-batch variations. At the same time, a wide variety of non-protein peroxidase mimics are now available that show strong potential to replace protein enzymes. Here, we review and analyze non-protein biosensors that represent a nexus of these two concepts: aptamer-based sensors (aptasensors) with optical detection (colorimetric, luminescent, or fluorescent) based on different peroxidase mimics, such as DNAzymes, nanoparticles, or metal-organic frameworks.
Topics: Peroxidase; Prospective Studies; Reproducibility of Results; Peroxidases; Oligonucleotides; Biosensing Techniques; Antibodies; Aptamers, Nucleotide
PubMed: 38275302
DOI: 10.3390/bios14010001 -
Proceedings of the National Academy of... Aug 2023Cells of vertebrate and invertebrate organisms express proteins specialized in membrane channel-based cell-cell communication that are absent in unicellular organisms....
Cells of vertebrate and invertebrate organisms express proteins specialized in membrane channel-based cell-cell communication that are absent in unicellular organisms. We recently described the prediction of some members of the large-pore channel family in kinetoplastids, consisting of proteins called unnexins, which share several structural features with innexin and pannexin proteins. Here, we demonstrated that the unnexin1 protein (Unx1) is delivered to the cell membrane, displaying a topology consisting of four transmembrane domains with C and N termini on the cytoplasmic side and form large-pore channels that are permeable to small molecules. Low extracellular Ca/Mg levels or extracellular alkalinization, but not mechanical stretching, increases channel activity. The Unx1 channel mediates the influx of Ca and does not form intercellular dye coupling between HeLa Unx1 transfected cells. Unx1 channel function was further evidenced by its ability to mediate ionic currents when expressed in oocytes. Downregulation of Unx1 mRNA with morpholine contains invasion. Phylogenetic analysis revealed the presence of Unx1 homologs in other protozoan parasites, suggesting a conserved function for these channel parasites in other protists. Our data demonstrate that Unx1 forms large-pore membrane channels, which may serve as a diffusional pathway for ions and small molecules that are likely to be metabolic substrates or waste products, and signaling autocrine and paracrine molecules that could be involved in cell invasion. As morpholinos-induced downregulation of Unx1 reduces the infectivity of trypomastigotes, the Unx1 channels might be an attractive target for developing trypanocide drugs.
Topics: Protein Subunits; Phylogeny; Cell Membrane; Cytoplasm; Morpholinos
PubMed: 37487087
DOI: 10.1073/pnas.2307898120 -
Chembiochem : a European Journal of... Jul 2024Cyclic dinucleotides (CDNs) have garnered popularity over the last decade as immunotherapeutic agents, which activate the cyclic GMP-AMP synthase-stimulator of...
Cyclic dinucleotides (CDNs) have garnered popularity over the last decade as immunotherapeutic agents, which activate the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway to trigger an immune response. Many analogs of 2'3'-cGAMP, c-di-GMP, and c-di-AMP have been developed and shown as effective cancer vaccines and immunomodulators for the induction of both the adaptive and innate immune systems. Unfortunately, the effectiveness of these CDNs is limited by their chemical and enzymatic instability. We recently introduced 5'-endo-phosphorothoiate 2'3'-cGAMP analogs as potent STING agonist with improved resistance to cleavage by clinically relevant phosphodiesterases. We herein report the synthesis of locked nucleic acid-functionalized (LNA) endo-S-CDNs and evaluate their ability to activate STING in THP1 monocytes. Interestingly, some of our synthesized LNA 3'3'-endo-S-CDNs can moderately activate hSTING REF haplotype (R232H), which exhibit diminished response to both 2'3'-cGAMP and ADU-S100. Also, we show that one of our most potent endo-S-CDNs has remarkable chemical (oxidants I and HO) and phosphodiesterase stability.
Topics: Membrane Proteins; Humans; Oligonucleotides; Nucleotides, Cyclic; THP-1 Cells
PubMed: 38720428
DOI: 10.1002/cbic.202400321 -
Bioanalysis Dec 2023Accurate detection and monitoring of therapeutic drug levels are vital for effective patient care and treatment management. Aptamers, composed of single-stranded DNA or... (Review)
Review
Accurate detection and monitoring of therapeutic drug levels are vital for effective patient care and treatment management. Aptamers, composed of single-stranded DNA or RNA molecules, are integral components of biosensors designed for both qualitative and quantitative detection of biological samples. Aptasensors play crucial roles in target identification, validation, detection of drug-target interactions and screening potential of drug candidates. This review focuses on the pivotal role of aptasensors in early disease detection, particularly in identifying biomarkers associated with various diseases such as cancer, infectious diseases and cardiovascular disorders. Aptasensors have demonstrated exceptional potential in enhancing disease diagnostics and monitoring therapeutic drug levels. Aptamer-based biosensors represent a transformative technology in the field of healthcare, enabling precise diagnostics, drug monitoring and disease detection.
Topics: Humans; Molecular Probes; Drug Monitoring; Aptamers, Nucleotide; Biomarkers; Neoplasms; Biosensing Techniques
PubMed: 37847048
DOI: 10.4155/bio-2023-0141 -
Journal of Inorganic Biochemistry Oct 2023Two oligonucleotide conjugates sharing the same sequence but incorporating a different 5'-terminal organometallic moiety were synthesized, by either direct mercuration...
Two oligonucleotide conjugates sharing the same sequence but incorporating a different 5'-terminal organometallic moiety were synthesized, by either direct mercuration in solution or oximation with an organomercury aldehyde on solid support. The potential of these conjugates to serve as new type of artificial ribonucleases was tested with a complementary 2´-O-methyl-RNA target sequence featuring a single cleavable RNA phosphodiester linkage. Both organomercury oligonucleotides greatly outperformed their metal-free counterparts as well as the previously reported small molecule organomercury RNA cleaving agent in catalytic activity, providing an important proof-of-concept. Compared to state-of-the-art metal-dependent artificial ribonucleases, however, the observed activity was modest.
Topics: Aldehydes; Oligonucleotides; RNA; Ribonucleases
PubMed: 37480764
DOI: 10.1016/j.jinorgbio.2023.112331 -
Progress in Molecular Biology and... 2024The prevalence of metabolic disorders is increasing exponentially and has recently reached epidemic levels. Over the decades, a large number of therapeutic options have...
The prevalence of metabolic disorders is increasing exponentially and has recently reached epidemic levels. Over the decades, a large number of therapeutic options have been proposed to manage these diseases but still show several limitations. In this circumstance, RNA therapeutics have rapidly emerged as a new hope for patients with metabolic diseases. 57 years have elapsed from the discovery of mRNA, a large number of RNA-based drug candidates have been evaluated for their therapeutic effectiveness and clinical safety under clinical studies. To date, there are seven RNA drugs for treating metabolic disorders receiving official approval and entering the global market. Their targets include hereditary transthyretin-mediated amyloidosis (hATTR), familial chylomicronemia syndrome, acute hepatic porphyria, primary hyperoxaluria type 1 and hypercholesterolemia, which are all related to liver proteins. All of these seven RNA drugs are antisense oligonucleotides (ASO) and small interfering RNA (siRNA). These two types of treatment are both based on oligonucleotides complementary to target RNA through Watson-Crick base-pairing, but their mechanisms of action include different nucleases. Such treatments show greatest potential among all types of RNA therapeutics due to consecutive achievements in chemical modifications. Another method, mRNA therapeutics also promise a brighter future for patients with a handful of drug candidates currently under development.
Topics: Humans; Oligonucleotides; Oligonucleotides, Antisense; RNA, Small Interfering; RNA, Messenger; Amyloid Neuropathies, Familial
PubMed: 38359998
DOI: 10.1016/bs.pmbts.2023.12.014 -
Cell Reports Methods Dec 2023The process of nucleic acid aptamer selection can be quite laborious and fraught with artifacts. In a work published in Nature Biotechnology, Singh et al. describe an...
The process of nucleic acid aptamer selection can be quite laborious and fraught with artifacts. In a work published in Nature Biotechnology, Singh et al. describe an approach that should allow more facile aptamer selection.
Topics: Aptamers, Nucleotide; Porosity; SELEX Aptamer Technique; Biotechnology; Biocompatible Materials; Hydrogels
PubMed: 38065094
DOI: 10.1016/j.crmeth.2023.100667 -
Molecules (Basel, Switzerland) Mar 2024Aptamers developed using in vitro Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technology are single-stranded nucleic acids 10-100 nucleotides in... (Review)
Review
Aptamers developed using in vitro Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technology are single-stranded nucleic acids 10-100 nucleotides in length. Their targets, often with specificity and high affinity, range from ions and small molecules to proteins and other biological molecules as well as larger systems, including cells, tissues, and animals. Aptamers often rival conventional antibodies with improved performance, due to aptamers' unique biophysical and biochemical properties, including small size, synthetic accessibility, facile modification, low production cost, and low immunogenicity. Therefore, there is sustained interest in engineering and adapting aptamers for many applications, including diagnostics and therapeutics. Recently, aptamers have shown promise as early diagnostic biomarkers and in precision medicine for neurodegenerative and neurological diseases. Here, we critically review neuro-targeting aptamers and their potential applications in neuroscience research, neuro-diagnostics, and neuro-medicine. We also discuss challenges that must be overcome, including delivery across the blood-brain barrier, increased affinity, and improved in vivo stability and in vivo pharmacokinetic properties.
Topics: Animals; Aptamers, Nucleotide; SELEX Aptamer Technique; Neurosciences; Antibodies; Ligands
PubMed: 38474636
DOI: 10.3390/molecules29051124 -
Biosensors & Bioelectronics Oct 2023Following the recent pandemic and with the emergence of cell-free nucleic acids in liquid biopsies as promising biomarkers for a broad range of pathologies, there is an... (Review)
Review
Following the recent pandemic and with the emergence of cell-free nucleic acids in liquid biopsies as promising biomarkers for a broad range of pathologies, there is an increasing demand for a new generation of nucleic acid tests, with a particular focus on cost-effective, highly sensitive and specific biosensors. Easily miniaturized electrochemical sensors show the greatest promise and most typically rely on the chemical functionalization of conductive materials or electrodes with sequence-specific hybridization probes made of standard oligonucleotides (DNA or RNA) or synthetic analogues (e.g. Peptide Nucleic Acids or PNAs). The robustness of such sensors is mostly influenced by the ability to control the density and orientation of the probe at the surface of the electrode, making the chemistry used for this immobilization a key parameter. This exhaustive review will cover the various strategies to immobilize nucleic acid probes onto different solid electrode materials. Both physical and chemical immobilization techniques will be presented. Their applicability to specific electrode materials and surfaces will also be discussed as well as strategies for passivation of the electrode surface as a way of preventing electrode fouling and reducing nonspecific binding.
Topics: Nucleic Acids; Biosensing Techniques; DNA; Peptide Nucleic Acids; Oligonucleotides; Electrodes; Electrochemical Techniques; Nucleic Acid Hybridization
PubMed: 37406480
DOI: 10.1016/j.bios.2023.115440