Novel poly(azoamide triazole)s containing twin azobenzene units in the backbone. Synthesis, characterization, and in vitro degradation studies

We describe the synthesis and characterization of four new light and reduction sensitives poly(azoamide triazole)s, in which the azobenzene units are found along the main chain of the macromolecule. These polymers were prepared by the azide-alkyne cycloaddition reaction catalyzed with copper (I) (CuAAC). They were obtained in high yield and with apparent molecular weights in the range from 95 to 148 kDa. All poly(azoamide triazole)s are soluble in polar aprotic solvents, and two of them are also soluble in chloroform showing good coating and film-forming properties. They were characterized by Fourier transform infrared, nuclear magnetic resonance (NMR), ultraviolet-visible spectroscopy and gel permeation chromatography (GPC). The photoisomerization study of the synthesized polymers has been carried out by UV–Vis spectroscopy, as well as their trans-cis-trans reversibility behavior. Differential scanning calorimetry (DSC) and themogravimetric analysis (TGA) were used to investigate their thermal properties. Results show that the polymers were amorphous and stable up to 300 °C under nitrogen. The hydrolytic degradation of films of these polymers has been studied in vitro under various conditions of pH and temperature and was monitored by GPC. Furthermore, the presence of azo units along the polymer backbone as cleavable groups provides access to their degradation by reduction. In this sense, the degradation of polymers has also been studied using sodium dithionite as a mimic of the enzyme azoreductase. The results of these studies show that the polymers are stable enough under hydrolytic physiological conditions, but they degrade rapidly when sodium dithionite is used. A preliminary study of biocompatibility of polymers PAAT1 and PAAT4 has been carried out. A hemolysis study with human red blood cells (hRBC) and a cytotoxicity study with human gingival fibroblasts (HGnF) have been carried out. The results obtained suggest that these polymers could be good candidates to be used as drug coating materials.Ministerio de Economía y Competitividad MAT2016-77345-C3-2-PUniversidad de Sevilla VI Plan Propio, PP2021/0000065

Engineering plasmonic nanomaterials for SERS applications

En conseqüència de les seves extraordinàries propietats, les nanopartícules d'or i plata s'han utilitzat en una varietat d'aplicacions, incloses les relacionades amb surface-enhanced Raman scattering (SERS) spectroscopy. La creixent ocupació d'aquesta tècnica com a eina analítica, va empènyer la investigació al studio de nous substrats SERS. Considerant aquestes dades, la thesis proposada té com a objectiu examinar estratègies per al disseny de nanomaterials nous i expandeix la molecular library de lligands de superfície per funcionalitzar les plataformes plasmòniques amb un enfocament especial a l'aplicació SERS. Primer, es proposa un mètode per a la formació homogènia d'una capa de plata sobre nuclis d'or amb diferents característiques geomètriques a través d'un procés de cold nanowelding. solució. En particular, la investigació realitzada ha demostrat la forta implicació d'alguns paràmetres experimentals en la generació d'estructures bimetàl·liques d'Au/Ag que conserven la morfologia del substrat d'or original. Segon, un nou derivat de azobenzè va ser proposat com a quimioreceptor eficaç i de baix cost per a la detecció SERS de tiols de baix pes molecular clínicament rellevants. Més concretament, els agregats de nanopartícules de plata col·loïdalment estables es van equipar amb el mercapte-azobenzè derivat que, en presència de biotíols, mostra importants alteracions espectrals SERS correlacionades quantitativament amb el trencament de l'enllaç diazo intervingut per molècules petites tiolades. Finalment es va estudiar la producció d'una graded-index (GRIN) lens que es va obtenir mitjançant l'acoblament jeràrquic de nanopartícules d'or en un nucli micromètric de sílice amb l'objectiu d'examinar les propietats exòtiques òptiques relacionades amb la complexa arquitectura i que consisteixen en la capacitat del sistema de concentrar la llum al centre de la microesfera.Debido a sus extraordinarias propiedades, las nanopartículas de oro y plata se han utilizado en una variedad de aplicaciones, incluidas las que estan relacionada con surface-enhanced Raman scattering (SERS) spectroscopy. El creciente empleo de esta técnica como herramienta analítica, empujó la investigación en el studio de nuevos sustratos SERS. Considerando estos datos, la thesis propuesta tiene como objetivo examinar estrategias para el diseño de nanomateriales novedosos y expande la molecular library de ligandos de superficie para funcionalizar las plataformas plasmónicas con un enfoque especial en la aplicación SERS. Primero, se propone un metodo para la formación homogenea de una capa de plata sobre núcleos de oro con diferentes características geométricas a través de un proceso de cold nanowelding- La nueva estrategia sintetica pretende superar las limitaciones identificadas en las reducciones químicas convencionales de sales metálicas en solución. En particular, la investigación realizada ha demostrado la fuerte implicación de algunos parámetros experimentales en la generación de estructuras bimetálicas de Au/Ag que conservan la morfología del sustrato de oro original. Segundo, un nuevo derivado de azobenceno fue propuesto como quimiorreceptor eficaz y de bajo coste para la detección SERS de tioles de bajo peso molecular clínicamente relevantes. Más concretamente, los agregados de nanopartículas de plata coloidalmente estables se equiparon con el mercapto-azobenceno derivado que, en presencia de biotioles, muestra importantes alteraciones espectrales SERS correlacionadas cuantitativamente con la rotura del enlace diazo mediada por moléculas pequeñas tioladas. Finalmente se estudió la producción de una graded-index (GRIN) lens que se obtuvo mediante el ensamblaje jerárquico de nanopartículas de oro en un núcleo micrométrico de sílice con el objetivo de examinar las propiedades exóticas ópticas relacionada con la compleja arquitectura y que consisten en la capacidad del sistema de concentrar la luz en el centro de la microesfera.Due to their exceptional properties, gold and silver nanoparticles have been used in a variety of applications, including those related to surface-enhanced Raman scattering (SERS) spectroscopy. In this regard, the increasing employment of this spectroscopic technique as an ultrasensitive analytical tool in a wide range of application fields, encouraged research in further improvements in the SERS substrates engineering. With these considerations in mind, the proposed dissertation aims to examine approaches for the design of novelty nanomaterials and expands the molecular library surface ligands that can decorate the plasmonic platforms with a special focus on SERS application. First, the formation of a smooth silver layer onto gold cores with different geometrical features through a cold nanowelding process was proposed as a new synthetical approach that overcomes the issues identified in the conventional chemical reductions of metal salts in solution. In particular, the conducted research has demonstrated the strong implication of some experimental parameters in the generation of bimetallic Au/Ag structures that retain the original gold substrate morphology. Secondly, a novel azobenzene derivative was synthesized as an efficient low-cost chemoreceptor for the SERS detection of clinically relevant low molecular weight thiols. More concretely, colloidally stable silver nanoparticle clusters were equipped with the obtained mercapto-azobenzene probe that, in the presence of biothiols, displays dramatic SERS spectral alterations quantitatively correlated to the small thiolated molecules-mediated breakage of the diazo bond. The dissertation finally reports a study conducted on the production of a metallic metamaterial. The resulting hybrid graded-index (GRIN) lens was obtained through the hierarchical assembly of gold nanoparticles onto a micrometric core of silica with the aim to examine the exotic optical properties to concentrate the light at the center of the microsphere deriving from the complex architecture

DNA結合蛋白をラベル化及びDNA架橋反応性を持つ機能性オリゴヌクレオチドの開発

要約のみTohoku University永次史課

Use of styrylbenzothiazole photoswitches in drugs for spatiotemporal control over microtubule-dependent biology

Thousands of biological processes take place within a single cell, every second of its existence. These processes are meticulously orchestrated and highly regulated in order to guarantee proper functioning and homeostasis within the organism. Multiple proteins assemble to form highly sophisticated molecular machines and structures to carry out certain tasks at exact timepoints, such as the microtubule-based mitotic spindle apparatus, which separates the chromosomes during mitosis; or the replisome that performs DNA replication with an unprecedented level of precision and reliability. The visualization of cells, their biological processes, and our understanding of the biomolecular function of proteins, have grown rapidly with the invention of groundbreaking new technologies and continuous improvement of existing instruments. Cryo electron microscopy, high-speed or super-resolution microscopy have considerably pushed the limits of temporal and spatial resolution past the micron spatial scale and millisecond time scale, opening new frontiers in modern biology. Small molecule drugs which can modulate, inhibit or amplify these machineries and their functions, have been a key factor to explore the molecular dynamics of proteins and cellular systems. Consequently, while selective small molecule inhibitors for certain protein targets have garnered much interest in drug discovery, as promising drug candidates to treat diseases, they have also become crucial research tools in order to perturb and study protein and network function. However, the technological leap has created a discrepancy between modern technology and classic drugs when used as research tools. With high-precision instruments able to observe highly dynamic cellular systems, it is the current set of molecular tools available to researchers that lack the spatiotemporal precision to control these dynamic cellular systems on a submicron and millisecond scale. Influencing or controlling biological processes with classic small molecule inhibitors without spatiotemporal specificity is a poor method to investigate a finely tuned machine. Part One of this work will focus on the introduction of innovative molecular tools that allow simple control over a highly dynamic cellular system: the microtubule cytoskeleton. This is intended to optimally utilize, and in return further develop, state of the art imaging techniques and to expand biologists arsenal of molecular tools that can help to answer key questions in neuroscience, embryology, and cytoskeleton research, by using light as a non-invasive, high-precision, bioorthogonal regulator for biological application. I introduce the styrylbenzothiazole (SBT) photoswitch as a research tool in cell biology, with initial application to reliably enable fast and reversible in situ optical control over the microtubule cytoskeleton. To situate this application I will discuss the problems of current state-of-the-art photoswitchable microtubule destabilizers, in particular azobenzene based PST-1, and showcase the SBTubs as an SBT-based alternative. We confirm their biological utility as photoswitchable tubulin inhibitors, compare their (photo)chemical & metabolic robustness, and test their compatibility to common fluorescent imaging tags over the azobenzene scaffold. Part Two will focus on consolidating the newly introduced SBTs as a powerful alternative photoswitch scaffold with complementary features to azobenzenes. I perform an SAR study and identify two lead compounds SBTub2M and SBTubA4P as low nanomolar and water-soluble photoswitchable antimitotics that enable photocontrol over microtubule dynamics and structure in 3D systems and animal models (D. rerio, X. tropicalis), thus bringing in vivo photopharmacology one step closer to realization. Furthermore, I report the first photopharmacology study of a styrylthiazole (ST) photoswitch. In the final part of my thesis I will apply this ST photoswitch scaffold instead to address the taxane binding site, showing its applicability and general features. I design and synthesize the first ever reported photoswitchable epothilones (STEpos) and offer proof of principle of their light-dependent stabilization of microtubules. Taken together, this research makes a contribution towards (a) spatiotemporal control of microtubule research, helping antimitotic photopharmaceuticals advance beyond 2D cell culture and (b) reshaping the field of photopharmacology by introducing a focus on photoswitches like the SBTs which respond to biologists’ technological needs & capacities, which will contribute to new SBT-based photopharmaceuticals both in microtubule research and beyond

Tris(azidoethyl)amine Hydrochloride; a Versatile Reagent for Synthesis of Functionalized Dumbbell Oligodeoxynucleotides

Triazole-cross-linked oligodeoxynucleotides were synthesized using the Cu(I) catalyzed alkyne–azide cycloaddition with tris(azidoethyl)amine hydrochloride and oligodeoxynucleotides possessing <i>N</i>-3-(propargyl)thymidine at both the 3′- and 5′-termini. Further installation of a functional molecule to the dumbbell oligodeoxynucleotides was achieved by utilizing the remaining azide group