Glycan Sequencing Based on Glycosidase-Assisted Nanopore Sensing

Journal of the American Chemical Society, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 2, 2025

Nanopores are promising sensors for glycan analysis with the accurate identification of complex glycans laying foundation nanopore-based sequencing. However, their applicability toward continuous sequencing has not yet been demonstrated. Here, we present a proof-of-concept by combining nanopore technology glycosidase-hydrolyzing reactions. By continuously monitoring changes in characteristic current generated translocation hydrolysis products through nanopore, sequence can be accurately identified based on specificity glycosidases. With machine learning, improved accuracy to over 98%, allowing reliable determination consecutive building blocks and glycosidic linkages chains while reducing need operator expertise. This approach was validated real samples, calibrated using hydrophilic interaction chromatography-high-performance liquid chromatography (HILIC-HPLC) mass spectrometry (MS). We achieved ten units natural chains, which provided first evidence feasibility nanopore-glycosidase-compatible system Compared traditional methods, this strategy enhances efficiency 5-fold. Additionally, introduced concept 'inverse sequencing', focuses electrical signal rather than monosaccharide identification. eliminates reliance fingerprint libraries typically required putative 'forward hydrolysis' strategies. When challenges both inverse strategies' addressed, will pave way establishing at single-molecule level.

Language: Английский

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Recent Progress in Developing Extracellular Vesicles as Nanovehicles to Deliver Carbohydrate-Based Therapeutics and Vaccines

Vaccines, Journal Year: 2025, Volume and Issue: 13(3), P. 285 - 285

Published: March 7, 2025

Cell-derived, nanoscale extracellular vesicles (EVs) have emerged as promising tools in diagnostic, therapeutic, and vaccine applications. Their unique properties including the capability to encapsulate diverse molecular cargo well versatility surface functionalization make them ideal candidates for safe effective vehicles deliver a range of biomolecules gene editing cassettes, therapeutic proteins, glycans, glycoconjugate vaccines. In this review, we discuss recent advances development EVs derived from mammalian bacterial cells use delivery carbohydrate-based protein therapeutics We highlight key innovations EVs’ design, characterization, deployment treating diseases Alzheimer’s disease, infectious diseases, cancers. challenges their clinical translation provide perspectives future within biopharmaceutical research landscape.

Language: Английский

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Fine-Tuning the Active-Site Microenvironment of β-Galactosidase to Enhance the Synthesis Ability of Galactooligosaccharides while Minimizing the Impairment to Transglycosylation Activity

Journal of Agricultural and Food Chemistry, Journal Year: 2025, Volume and Issue: unknown

Published: April 22, 2025

Most reported mutations at -1 subsites of β-galactosidases that improved galactooligosaccharides (GOS) synthesis ability significantly reduced total activity (e.g., <10%), likely due to the introduction significant disturbances within active-site microenvironment. In this study, a fine-tuning strategy encompassing aromatic residue interchange as well substitution among T, S, and A was proposed subsequently evaluated in Bgal1-3 three commercial β-galactosidases. For each them, 2-4 positive mutants were acquired with residual activities 30-357%. When 40% (w/v) lactose employed substrate, their GOS yields 1.2-18% higher than those wild types. Moreover, best produced greater amounts skim milk (2.9-11.8 g/L higher) conversion rate 90%. Ultimately, mutation set (∼14 mutations) designed for convenience using approach other glycoside hydrolases. This may hold great potential promoting enzymatic valuable carbohydrate-containing compounds.

Language: Английский

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Photocrosslinking and capture for the analysis of carbohydrate-dependent interactions

Bioorganic & Medicinal Chemistry Letters, Journal Year: 2024, Volume and Issue: 117, P. 130077 - 130077

Published: Dec. 20, 2024

Language: Английский

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