Liyuan Liang

Aspects inorganiques de la chimie “click” dans les polymères et dendrimères : synthèses, stabilisation de nanoparticules et catalyse: La thèse concerne les aspects inorganiques de la réaction des alcynes terminaux avec les azotures, la principale des réactions dites ”click”. La catalyse de cette réaction par des dendrimères centrés sur le cuivre (I) nous a permis de mettre en évidence des effets dendritiques originaux. Les assemblages dendritiques et polymériques fonctionnels réalisés à l’aide cette réaction “click”, en particulier avec des carboranes, nous ont conduits à la stabilisation de nanoparticules d’or et de palladium à partirde cations coordonnés aux triazoles “click”. La catalyse très efficace dans aqueux milieu et dans les conditions ambiantes de formation de liaison carbone-carbone a été réalisée à l’aide de très faibles quantités de ces nanoparticulesde palladium stabilisées.

The COVID-19 pandemic caused by the SARS-CoV-2 virus has exposed the urgency of research on rapid and efficient virus detection and strategies to inhibit its replication. Previous studies have mostly focused on traditional immunoassay or optical methods, but they have limitations in terms of sensitivity, timeliness, and in-depth analysis of molecular interaction mechanisms. Solid-state nanopore single-molecule detection methods, which can monitor molecular conditions in real time at the single-molecule level, bring new opportunities to solve this problem. The nucleocapsid protein (N protein) of SARS-CoV-2 was systematically investigated under different conditions, such as external drive voltage, pH, nanopore size, and N protein concentration. The translocation of the N protein through the nanopore was then analyzed. Subsequently, we analyzed the translocation characteristics of the N protein, RNA, and N protein–RNA complexes. With the aid of EMSA experiments, we conclusively confirmed that RNA binds to the N protein. Building on this finding, we further explored small molecules that could affect the nanopore translocation of N protein–RNA complexes, such as gallocatechin gallate (GCG), epigallocatechin gallate (EGCG), and the influenza A viral inhibitor Nucleozin. The results show that GCG can disrupt the liquid-phase condensation of the N protein–RNA complex and inhibit the replication of the N protein. Meanwhile, the structural isomer EGCG of GCG and the small molecule Nucleozin can also block RNA-triggered N protein liquid–liquid phase separation (LLPS). Our results confirmed that GCG, EGCG, and Nucleozin exhibit antagonistic effects on the N protein, with differences in their effective concentrations and the potency of their antagonism. Herein, using solid-state nanopore single-molecule detection technology, we developed an experimental method that can effectively detect RNA-induced changes in N protein properties and the regulatory effects of small molecules on the LLPS of N protein–RNA complexes. These findings not only provide highly valuable insights for in-depth research on the molecular interactions involved in viral replication, but also open up promising new avenues for future responses to similar viral outbreaks, the development of a rapid and effective detection method based on solid-state nanopores and single-molecule detection, and antiviral therapies targeting N protein–RNA interactions.

Noncanonical quadruplexes (G4s) in the nucleic acids represent specific secondary structures that correlate and participate in important biological processes, including telomeric propagation and tumor cell proliferation, and are close to the life span. The interaction of G4 with specific proteins and monitoring of the unfolding process are important to understand the development and evolution of some diseases and for further regulation of telomere and to disclose the mechanism of typical cancers. Different from the most reported single-molecule tweezers’ manipulation, this work provides a nanopore-based electric approach for label-free monitoring of the unfolding process of both DNA- and RNA-G4 under various conditions. Twenty nanomolar of hTel sequence could be mostly unfolded via incubation with 10 nM of both TEP1 that is associated with telomerase and helicase RTEL1 under weak acidic conditions for 1 h, and this process could be recorded in a single-molecule nanopore device with a pore diameter of 3.7 nm in 0.5 M CsCl buffered solution at 150 mV. TEP1 is proved to specifically interplay with hybrid G4 and nearly does not unfold parallel G4. The efficient and selective unfolding process of RNA-G4 originated from SARS-CoV by helicase nsp13 is also demonstrated at pH 5 in 2 M LiCl with a 3.6-nm pore; with 50% molar ratio of helicase nsp13, RNA-1574-G4 could be selectively and significantly unfolded in 1 h. This work presents the selective unfolding of both DNA- and RNA-G4 by specific proteins, which is a new approach to modulate and monitor the secondary structure of nucleic acids, and shed light on the understanding of the mechanism of the interaction between nucleic acids and proteins and the related kinetics.

Oligopeptides in anti-aging cosmetics stand out as active ingredients to interact with skin cell and accelerate the collagen synthesis and fibroblast proliferation. Some of them act as neurotransmitter- or enzyme inhibitor, while others are signal or carrier peptides. The ensemble techniques for the extraction and analysis of the bioactive peptides in cosmetic production involve ultrafiltration, enzymatic hydrolysis, fermentation, and high-performance liquid chromatography. This work provides a single-molecule approach for the resolution of various typical oligopeptides in anti-aging cosmetics. A nanopore with an aperture diameter of ~ 2 nm is efficient for the shortest tripeptide to achieve good signal to noise ratio and translocation frequency. Oligoeptides with three to eight amino acids could be discriminated with a ~ 2 nm single SiN_x nanopore, this is also proved by AI modeling with an accuracy of ~ 90%, except the argireline and hexa-peptide-9 that hold similar translocation behavior in KCl. The assay for the argireline in three commercial anti-aging cosmetics reveals that the product from Viribati is purer than the other two and harbors the majority of argireline according to the comparison setups in a single nanopore device. Our work provides a significant insight in the qualification and quality control in anti-aging cosmetic market.