Biology subjects

Nguyen, A.-T.

Publications and source records attributed to Nguyen, A.-T..

6 recordsLinked to original sources

bioRxiv2026

Flow alters fibrin molecular and network structure and decreases binding of fibrinolytic enzymes

Thrombolysis with tissue Plasminogen Activator (tPA), approved for treating acute ischemic stroke (AIS) within 3-4.5 h of symptom onset, converts Plasminogen (Plg) to Plasmin to degrade fibrin, while fibrin also enhances Plg activation by binding to both tPA and Plg. Therefore, in this study, we investigated whether arterial-like flow, characteristic of AIS, alters fibrin structure and susceptibility to fibrinolysis. Using a newly developed platform for quantitative imaging and spectroscopy, we found that flow generates denser fibrin networks with reduced molecular transport despite reduced protofibril packing within fibrin fibers. Raman spectroscopy revealed an -helix-to-{beta}-sheet transition, accompanied by reduced Plg and tPA binding, although the reduction in tPA binding emerged only after prolonged flow exposure. Consistently, multi-scale molecular dynamics simulations showed that the Plg binding site destabilized at lower forces than the primary tPA binding site. Together, these multi-scale findings help explain the limited efficacy and narrow therapeutic window of thrombolysis.

biophysics↗

bioRxiv2026

A High-Throughput Platform for Rapidly Adapting DNA Aptamers to SARS-CoV-2 Evolution

Rapid pathogen evolution threatens public health by eroding the effectiveness of vaccines, therapeutics, and diagnostic tools. Although spike protein targeting monoclonal antibodies (mAbs) were developed within 10-12 months of the initial outbreak to serve as key theranostic agents, their redesign has struggled to keep pace with viral evolution, rendering many neutralizing antibodies ineffective. Here we demonstrate a novel platform that combines a random-rational hybrid library diversification with high-throughput MiSeq screening to rapidly reprogram aptamers against emerging SARS-CoV-2 spike variants. Interactions between 3 different spike proteins and 11,806 unique aptamer variant designs were profiled within a few days. Starting from a 40-nt aptamer originally selected against wild-type (WT) spike protein, our screen identified a Delta-binding mutant with a 4-fold affinity improvement and an Omicron-binding mutant that converted undetectable binding into nanomolar affinity. We also identified a WT-selective mutant with substantially reduced affinity for Delta, as well as previously unrecognized bases that critically contribute to spike recognition. Integrating high-throughput binding data with molecular dynamics simulations further revealed sequence-dependent structural features underlying variant-specific aptamer-spike interactions. Finally, we developed a sensor based on the identified WT-selective aptamer mutant, enabling highly specific detection of the WT spike protein with robust performance. Together, this work establishes a rapid and adaptable aptamer engineering platform for rapid adaptation of aptamers to evolving pathogens in future pandemics.

bioengineering↗

bioRxiv2026

Lifetime-based multiplexed detection of viral RNA using fluorogenic aptamers

Fluorogenic aptamers (FAPs) are emerging molecular probes for viral RNA and DNA sensing. However, their use in multiplexed nucleic acid sensing has been hindered by cross-reactivity and overlapping emission spectra. Here we address these limitations by introducing a fluorescence-lifetime-based multiplexed detection strategy using variants of the DNA fluorogenic aptamer Lettuce that exhibits distinct fluorescence lifetimes when complexed with the fluorogen TO1-biotin. To effectively evolve Lettuce for diverse lifetimes, we developed a large-scale screening platform, termed FAP-FLIM-NGS (fluorogenic aptamer-based fluorescence lifetime imaging microscopy on next-generation sequencing chips), which measures the fluorescence lifetimes of [~]104 Lettuce/TO1-biotin complexes directly on an Illumina MiSeq flow cell. Using this approach, three variants with markedly different lifetimes were identified: a single mutant (smC14T, 6.0 ns) and two double mutants (dmA5T/C14T, 5.2 ns, and dmA5T/T22A, 4.4 ns). To demonstrate the utility of these Lettuce variants in multiplexed detection, a set of split Lettuce probes targeting viral RNA fragments derived from SARS-CoV-2, MERS-CoV, and influenza A were designed and tested. Phasor plot analysis confirmed that these probes can robustly distinguish individual targets as well as mixtures containing any two or all three targets purely based on distinct fluorescence lifetimes of probes, thereby overcoming the challenges of cross-reactivity and spectral overlap. Beyond this proof of concept, our findings establish a generalizable strategy for engineering FAPs with customized photophysical properties, opening new avenues for next-generation diagnostics and molecular sensing technologies.

bioengineering↗

bioRxiv2026

Mechanistic insights into the color transformation of a non-FRET substrate for RNase activity detection

DNA-templated silver nanoclusters (DNA/AgNCs) have created a new class of non-FRET DNase substrates, termed Subak, that exhibits a color change upon DNase digestion. Although Subak substrates offer advantages such as ratiometric readouts and low manufacturing costs over traditional FRET substrates, the mechanism governing AgNC color switching remains unclear. Here, using a site-specific cleavage strategy, we identify color-switching hotspots and demonstrate that AgNC transformation can be controlled by the cleavage positions within the nucleic acid host. Our data support a cleavage-driven reorganization of the AgNC coordination environment, converting a non-emissive precursor into a red-emitting cluster, rather than direct enzyme-cluster interactions. Leveraging this insight, we engineer rSubak, an RNA-incorporated Subak that displays 95 nm red shift (530 to 625 nm) upon RNase cleavage. In amplification-free CRISPR/Cas13 assays for SARS-CoV-2, influenza A (A/H5N1), and measles viruses (MV) detection, rSubak achieved a limit of detection of 0.3 pM, superior to that of the commercial RNaseAlert ([~]250 pM). Collectively, our results establish Subak as a generalizable, non-FRET platform for sensitive ratiometric reporting the activities of diverse nucleases.

bioengineering↗

bioRxiv2025

Differential Glutamatergic Inputs to Semilunar Granule Cells and Granule Cells Underscore Dentate Gyrus Projection Neuron Diversity

Semilunar Granule Cells (SGCs) are sparse dentate gyrus projection neurons whose role in the dentate circuit, including pathway specific inputs, remains unknown. We report that SGCs receive more frequent spontaneous excitatory synaptic inputs than granule cells (GCs). Dual GC-SGC recordings identified that SGCs receive stronger medial entorhinal cortex and associational synaptic drive but lack short-term facilitation of lateral entorhinal cortex inputs observed in GCs. SGCs dendritic spine density in proximal and middle dendrites was greater than in GCs. However, the strength of commissural inputs and dendritic input integration, examined in passive morphometric simulations, were not different between cell types. Activity dependent labeling identified an overrepresentation of SGCs among neuronal ensembles in both mice trained in a spatial memory task and task naive controls. The divergence of modality specific inputs to SGCs and GCs can enable parallel processing of information streams and expand the computational capacity of the dentate gyrus.

neuroscience↗

bioRxiv2024

Fluorogenic Aptamer Optimizations on a Massively Parallel Sequencing Platform

Fluorogenic aptamers (FAPs) have become an increasingly important tool in cellular sensing and pathogen diagnostics. However, fine-tuning FAPs for enhanced performance remains challenging even with the structural details provided by X-ray crystallography. Here we present a novel approach to optimize a DNA-based FAP (D-FAP), Lettuce, on repurposed Illumina next-generation sequencing (NGS) chips. When substituting its cognate chromophore, DFHBI-1T, with TO1-biotin, Lettuce not only shows a red-shifted emission peak by 53 nm (from 505 to 558 nm), but also a 4-fold bulk fluorescence enhancement. After screening 8,821 Lettuce variants complexed with TO1-biotin, the C14T mutation is found to exhibit an improved apparent dissociated constant ([Formula] vs. 0.82 {micro}M), an increased quantum yield (QY: 0.62 vs. 0.59) and an elongated fluorescence lifetime ({tau}: 6.00 vs. 5.77 ns), giving 45% more ensemble fluorescence than the canonical Lettuce/TO1-biotin complex. Molecular dynamic simulations further indicate that the {pi}-{pi} stacking interaction is key to determining the coordination structure of TO1-biotin in Lettuce. Our screening-and-simulation pipeline can effectively optimize FAPs without any prior structural knowledge of the canonical FAP/chromophore complexes, providing not only improved molecular probes for fluorescence sensing but also insights into aptamer-chromophore interactions.

biophysics↗