Chiral Matching between Nucleic Acids and Polypeptides Facilitates Liquid-Liquid Phase Separation.

Liquid-liquid phase separation (LLPS) is a newly discovered phenomenon to modulate a multitude of cellular functions. Despite its importance, the full evolution mechanism of LLPS starting from intramolecular interactions to intermolecular condensations has yet to be revealed. In this study, we investigated a representative LLPS formed between negatively charged nucleic acids poly(G-quadruplex) and positively charged peptides poly(lysine).

Fluorogenic Linkage Integration for Nonfluorescent Transformations (FLINT).

Since its creation, single-molecule optical imaging has revolutionized the study of catalytic processes, yet its application largely relies on probing fluorogenic reactions. To overcome this limitation, we propose the Fluorogenic Linkage Integration for Nonfluorescent Transformation (FLINT) approach, an imaging method to resolve nonfluorogenic reactions at the single-molecule level. Using glucose oxidation as a model reaction, we coupled this nonfluorogenic reaction with a fluorogenic Amplex Red (AR) → resorufin (RF) transformation to create a cascading reaction.

design of a mechano-pharmaceutical screening platform against formation of individual beta-amyloid oligomers.

Small molecules that can reduce the neurotoxic beta-amyloid (Aβ) aggregates in the brain provide a potential treatment for Alzheimer disease (AD). Most screening methods for small-molecule hits focus on the overall Aβ aggregations without a specific target, such as the very first association step (i.e.

Decoupling Activity and Specificity in Coronazymes.

Specificity and activity are often at odds for natural enzymes. In this work, specificity and activity in coronazymes made of an Au nanoparticle (AuNP) and coated with DNA aptamer for glucose substrates are decoupled. By single-molecule fluorescent MT-HILO (magnetic tweezers coupled with highly inclined and laminated optical sheet) microscopy, it is found that this coronazyme has ≈30 times higher activity on the d-glucose compared to bare AuNP nanozymes.

Electroanalytical Quantification of DNA Chirality.

Although chirality is critical for molecular properties and functions, experimental quantification of chirality is lacking. Herein, we performed cyclic voltammetry (CV) under polarized magnetic fields to provide a unified scale to quantify and compare DNA chirality. We observed the largest electron spin polarization in DNA structures with opposite chiral senses, which is consistent with the effect of chiral-induced spin selectivity (CISS).

Catalytic Relaxation of Kinetically Trapped Intermediates by DNA Chaperones.

Common in biomacromolecules, kinetically trapped misfolded intermediates are often detrimental to the structures, properties, or functions of proteins or nucleic acids. Nature employs chaperone proteins but not nucleic acids to escort intermediates to correct conformations. Herein, we constructed a Jablonski-like diagram of a mechanochemical cycle in which individual DNA hairpins were mechanically unfolded to high-energy states, misfolded into kinetically trapped states, and catalytically relaxed back to ground-state hairpins by a DNA chaperone.

Dynamic Structures and Fast Transition Kinetics of Oxidized G-Quadruplexes.

8-oxoguanines (8-oxoG) in cells form compromised G-quadruplexes (GQs), which may vary GQ mediated gene regulations. By mimicking molecularly crowded cellular environment using 40% DMSO or sucrose, here it is found that oxidized human telomeric GQs have stabilities close to the wild-type (WT) GQs. Surprisingly, while WT GQs show negative formation cooperativity between a Pt(II) binder and molecularly crowded environment, positive cooperativity is observed for oxidized GQ formation.

Methylene blue accelerates liquid-to-gel transition of tau condensates impacting tau function and pathology.

Preventing tau aggregation is a potential therapeutic strategy in Alzheimer's disease and other tauopathies. Recently, liquid-liquid phase separation has been found to facilitate the formation of pathogenic tau conformations and fibrillar aggregates, although many aspects of the conformational transitions of tau during the phase transition process remain unknown. Here, we demonstrate that the tau aggregation inhibitor methylene blue promotes tau liquid-liquid phase separation and accelerates the liquid-to-gel transition of tau droplets independent of the redox activity of methylene blue.

Single-Molecular Dissection of Liquid-Liquid Phase Transitions.

Interaction between peptides and nucleic acids is a ubiquitous process that drives many cellular functions, such as replications, transcriptions, and translations. Recently, this interaction has been found in liquid-liquid phase separation (LLPS), a process responsible for the formation of newly discovered membraneless organelles with a variety of biological functions inside cells. In this work, we studied the molecular interaction between the poly-l-lysine (PLL) peptide and nucleic acids during the early stage of an LLPS process at the single-molecule level using optical tweezers.

Ensemble Force Spectroscopy of a G-Quadruplex Cluster on a Single-Molecule Platform.

Single-molecule methods offer high sensitivities with precisions superior to bulk assays. However, these methods are low in throughput and cannot repetitively interrogate the same cluster of molecular units. In this work, we investigate a tandem array of G-quadruplexes on a single-molecule DNA template with a throughput of at least two orders of magnitude higher than single-molecule force spectroscopy.

Mechanical Stability of DNA Corona Phase on Gold Nanospheres.

Noncovalent adsorption of biopolymers on the surface of gold nanoparticles (AuNPs) forms a corona phase that drastically diversify AuNP functions. However, mechanical stabilities of such corona phase are still obscure, hindering the application of biopolymer-coated AuNPs. Here, using optical tweezers, we have observed, for the first time, that DNA corona phase adsorbed on a 5 nm AuNP via two (dA) strands in proximity can withstand an average desorption force of 40 pN, which is higher than the stall force of DNA/RNA polymerases.

Ensemble Force Spectroscopy by Shear Forces.

Single-molecule techniques based on fluorescence and mechanochemical principles provide superior sensitivity in biological sensing. However, due to the lack of high throughput capabilities, the application of these techniques is limited in biophysics. Ensemble force spectroscopy (EFS) has demonstrated high throughput in the investigation of a massive set of molecular structures by converting mechanochemical studies of individual molecules into those of molecular ensembles.

Single-molecule displacement assay reveals strong binding of polyvalent dendrimer ligands to telomeric G-quadruplex.

Binding between a ligand and a receptor is a fundamental step in many natural or synthetic processes. In biosensing, a tight binding with a small dissociation constant (K) between the probe and analyte can lead to superior specificity and sensitivity. Owing to their capability of evaluating competitors, displacement assays have been used to estimate K at the ensemble average level.

Small Molecules Modulate Liquid-to-Solid Transitions in Phase-Separated Tau Condensates.

In Tau protein condensates formed by the Liquid-Liquid Phase Separation (LLPS) process, liquid-to-solid transitions lead to the formation of fibrils implicated in Alzheimer's disease. Here, by tracking two contacting Tau-rich droplets using a simple and nonintrusive video microscopy, we found that the halftime of the liquid-to-solid transition in the Tau condensate is affected by the Hofmeister series according to the solvation energy of anions. After dissecting functional groups of physiologically relevant small molecules using a multivariate approach, we found that charged groups facilitate the liquid-to-solid transition in a manner similar to the Hofmeister effect, whereas hydrophobic alkyl chains and aromatic rings inhibit the transition.

Machine-Learning-Assisted Analysis of Colorimetric Assays on Paper Analytical Devices.

Paper-based analytical devices (PADs) employing colorimetric detection and smartphone images have gained wider acceptance in a variety of measurement applications. PADs are primarily meant to be used in field settings where assay and imaging conditions greatly vary, resulting in less accurate results. Recently, machine-learning (ML)-assisted models have been used in image analysis.

Mechanical unfolding of ensemble biomolecular structures by shear force.

Mechanical unfolding of biomolecular structures has been exclusively performed at the single-molecule level by single-molecule force spectroscopy (SMFS) techniques. Here we transformed sophisticated mechanical investigations on individual molecules into a simple platform suitable for molecular ensembles. By using shear flow inside a homogenizer tip, DNA secondary structures such as i-motifs are unfolded by shear force up to 50 pN at a 77 796 s shear rate.

Detecting the Coronavirus (COVID-19).

The COVID-19 pandemic has created huge damage to society and brought panic around the world. Such panic can be ascribed to the seemingly deceptive features of COVID-19: Compared to other deadly viral outbreaks, it has medium transmission and mortality rates. As a result, the severity of the causative coronavirus, SARS-CoV-2, was deeply underestimated by society at the beginning of the COVID-19 outbreak.

Selection of appropriate protein assay method for a paper microfluidics platform.

Background: Paper-analytical devices (PADs) have gained popularity as a simple and low-cost alternative for determining a wide range of analytes including proteins. Even though several colorimetric PADs methods for protein estimation are reported in literature, they lack justification for the chosen method and parameters therein.

Aim: Major aim of this work was to thoroughly evaluate the most commonly used colorimetric protein assays and recommend the most appropriate method for PADs platform.

Duplex DNA Is Weakened in Nanoconfinement.

For proteins and DNA secondary structures such as G-quadruplexes and i-motifs, nanoconfinement can facilitate their folding and increase structural stabilities. However, the properties of the physiologically prevalent B-DNA duplex have not been elucidated inside the nanocavity. Using a 17-bp DNA duplex in the form of a hairpin stem, here, we probed folding and unfolding transitions of the hairpin DNA duplex inside a DNA origami nanocavity.

Evaluation of secondary metabolites, antioxidant activity, and color parameters of Nepali wines.

We evaluated the quality of wines produced in Nepal in terms of phenolic, flavonoid, anthocyanin and tannin content, antioxidant capacity, and color parameters using spectrophotometric methods. The total phenolic content, total flavonoid content, and total antioxidant activities in Nepali wines ranged from 85.5 to 960.