The work presented here has a multidisciplinary character, having as a common factor the characterization of self-assembled nanostructures through force spectroscopy. Exploring AFM as a tool for characterizing self-assembly and interaction forces in soft matter nanostructures, three different Bio and nonbiological systems where investigated, all of them share the common characteristic of being soft matter molecular structures at the nanoscale. The studied systems in question are: a) Polyelectrolyte – lipid nanocomposites. Single polyelectrolyte adsorption-desorption from supported lipid bilayers, b) Polyelectrolyte brushes and c) Virus-Like particles (VLPs). The scientific interest and industrial applications for each of these different nanostructures is broad, and their potential uses in the near future ranges from smart nanocontainers for drug and gene delivery, surface platforms for molecular recognition to the development of new nanodevices with ultrasensitive external stimuli responsiveness. These nano-structures are constructed following assembly of smaller subunits and belong to representative examples of soft matter in modern nanotechnology. The stability, behavior, properties and long term durability of these self-organized structures depends strongly on the environmental conditions to which they are exposed since their building mechanism is a balance between attractive noncovalent interactions and momentum transmitted collisions due Brownian motion of the solvent molecules. For example a set of long chain molecules firmly attached to one end to a surface will alter their conformation as the space between them is reduced or the environmental conditions are modified (i.e. ionic strength, pH or temperature). For a highly packed condition, this fuzzy surface known as a polyelectrolyte brush will then behave as a responsive material with tunable responsiveness. Thus the objective in the present case was to investigate the change in morphology and the mechanical response of a polyelectrolyte brush to external forces by application of AFM nanoindentations under different ionic strength conditions. The degree of penetration of the AFM tip through the brush will provide insights into the forces exerted by the brush against the tip. Compressions on the brush should aid to characterize its changes in compressibility for different salt concentrations. For the second chosen system, the interaction between two assembled interfaces was investigated at the single molecular level. A multilayered film formed by the consecutive assembly of oppositely charged polyelectrolytes and subsequently coated with a lipid membrane represents a fascinating soft composite material resembling more than a few structural components emerging in living organisms. The fluid bilayer, thus provide a biocompatible interface where additional functionalities can further be integrated (fusion peptides for instance). The smooth polymer cushion confers not only structural flexibility but also adaptability of the chosen substrate properties to be coated. This type of interface could be useful in the development of novel molecular biosensors with single molecule recognition capacities or in the fabrication of assays against pathogenic agents. The aim of this project was to study the molecular binding mechanism between the last polyelectrolyte layer and the lipid head group of the lower lipid leaflet. Understanding this adsorption mechanism between both interfaces, should likewise contribute to improve the fabrication of lipid coated polymeric nano/micro capsules with targeting properties. For example this could be critical in the field of nonviral gene therapy, where the improvement in the design of condensates of nucleic acids and other polymers with lipids (lipoplexes) are of main interest for its posterior use as delivery vectors. Finally, viral capsids were investigated. These naturally occurring assembled nanocontainers within living organisms stand for a remarkable example of nature’s morphological designs. These structures self-assemble from a small number of different proteins occurring in identical copies. The capsid as a self-assembled structure carries multiple functions: compaction of the genome, protection against external chemical threats, target recognition, structural support and finally facilitating the release of the genome into the host cell. It is highly interesting how these different functions are organized within the capsid which consists, for example, in the case of the norovirus of 180 identical copies of one single protein. Therefore, the mechanical stability and elastic properties of virus-like particles of Rubella and Norovirus were investigated by external application of loading forces with an AFM tip. The measurements were performed under conditions relevant for the virus infection mechanism. The applied compressions on these protein shells at pH values mimicking the virus life cycle will aid to learn about possible internal transitions among proteins which may be important for switching between the various functions of the capsid. The choice of two unrelated viral systems with different entry pathways into the cell and with different morphological architectures is expected to reveal crucial information about the stability and mechanical resistance to deformation of these empty membrane-coated and bare viral capsids. This last might provide clues on the stage of particle disassembly and cargo release during the final step of the infection process.

No description available.

The work presented here has a multidisciplinary character, having as a common factor the characterization of self-assembled nanostructures through force spectroscopy. Exploring AFM as a tool for characterizing self-assembly and interaction forces in soft matter nanostructures, three different Bio and nonbiological systems where investigated, all of them share the common characteristic of being soft matter molecular structures at the nanoscale. The studied systems in question are: a) Polyelectrolyte – lipid nanocomposites. Single polyelectrolyte adsorption-desorption from supported lipid bilayers, b) Polyelectrolyte brushes and c) Virus-Like particles (VLPs). The scientific interest and industrial applications for each of these different nanostructures is broad, and their potential uses in the near future ranges from smart nanocontainers for drug and gene delivery, surface platforms for molecular recognition to the development of new nanodevices with ultrasensitive external stimuli responsiveness. These nano-structures are constructed following assembly of smaller subunits and belong to representative examples of soft matter in modern nanotechnology. The stability, behavior, properties and long term durability of these self-organized structures depends strongly on the environmental conditions to which they are exposed since their building mechanism is a balance between attractive noncovalent interactions and momentum transmitted collisions due Brownian motion of the solvent molecules. For example a set of long chain molecules firmly attached to one end to a surface will alter their conformation as the space between them is reduced or the environmental conditions are modified (i.e. ionic strength, pH or temperature). For a highly packed condition, this fuzzy surface known as a polyelectrolyte brush will then behave as a responsive material with tunable responsiveness. Thus the objective in the present case was to investigate the change in morphology and the mechanical response of a polyelectrolyte brush to external forces by application of AFM nanoindentations under different ionic strength conditions. The degree of penetration of the AFM tip through the brush will provide insights into the forces exerted by the brush against the tip. Compressions on the brush should aid to characterize its changes in compressibility for different salt concentrations. For the second chosen system, the interaction between two assembled interfaces was investigated at the single molecular level. A multilayered film formed by the consecutive assembly of oppositely charged polyelectrolytes and subsequently coated with a lipid membrane represents a fascinating soft composite material resembling more than a few structural components emerging in living organisms. The fluid bilayer, thus provide a biocompatible interface where additional functionalities can further be integrated (fusion peptides for instance). The smooth polymer cushion confers not only structural flexibility but also adaptability of the chosen substrate properties to be coated. This type of interface could be useful in the development of novel molecular biosensors with single molecule recognition capacities or in the fabrication of assays against pathogenic agents. The aim of this project was to study the molecular binding mechanism between the last polyelectrolyte layer and the lipid head group of the lower lipid leaflet. Understanding this adsorption mechanism between both interfaces, should likewise contribute to improve the fabrication of lipid coated polymeric nano/micro capsules with targeting properties. For example this could be critical in the field of nonviral gene therapy, where the improvement in the design of condensates of nucleic acids and other polymers with lipids (lipoplexes) are of main interest for its posterior use as delivery vectors. Finally, viral capsids were investigated. These naturally occurring assembled nanocontainers within living organisms stand for a remarkable example of nature’s morphological designs. These structures self-assemble from a small number of different proteins occurring in identical copies. The capsid as a self-assembled structure carries multiple functions: compaction of the genome, protection against external chemical threats, target recognition, structural support and finally facilitating the release of the genome into the host cell. It is highly interesting how these different functions are organized within the capsid which consists, for example, in the case of the norovirus of 180 identical copies of one single protein. Therefore, the mechanical stability and elastic properties of virus-like particles of Rubella and Norovirus were investigated by external application of loading forces with an AFM tip. The measurements were performed under conditions relevant for the virus infection mechanism. The applied compressions on these protein shells at pH values mimicking the virus life cycle will aid to learn about possible internal transitions among proteins which may be important for switching between the various functions of the capsid. The choice of two unrelated viral systems with different entry pathways into the cell and with different morphological architectures is expected to reveal crucial information about the stability and mechanical resistance to deformation of these empty membrane-coated and bare viral capsids. This last might provide clues on the stage of particle disassembly and cargo release during the final step of the infection process.

Dans ce manuscrit nous décrivons des études originales sur les propriétés photophysiques des nanotubes de carbone monoparois réalisées à l'échelle de la molécule unique. Nous nous sommes concentrés sur deux problématiques : l'effet du remplissage des nanotubes par de l'eau sur leurs propriétés de photoluminescence (PL) et la création de complexes multi-excitoniques en régime de forte excitation laser. Dans ce but nous avons utilisé une combinaison de microscopie, de spectroscopie et de mesures de déclin de PL. Nos résultats montrent pour des nanotubes de différents diamètres un décalage vers le rouge des énergies d'émission pour les nanotubes remplis d'eau. De plus, des déclins de PL biexponentiels sont obtenus pour des nanotubes individuels (6,4) vides et remplis d'eau. Les temps de déclin caractéristiques de ces deux espèces de nanotubes sont distincts, avec une réduction de la composante courte pour les nanotubes remplis. Ces résultats sont expliqués par une augmentation de la constante diélectrique dans les nanotubes remplis d'eau. Notre résultat le plus conséquent a été l'observation de la génération de trions dans des nanotubes non dopés en utilisant des moyen tout optiques. L'émission du trion apparaît dans les spectres de PL comme une bande latérale décalée vers le rouge. Basé sur nos observations expérimentales, nous proposons le modèle de génération du trion suivant lequel dans un régime multiexcitonique les interactions d'annihilation exciton-exciton créent des porteurs de charge qui sont piégés dans les fluctuations de potentiel électrostatique induites par les inhomogénéités de l'environnement. L'absorption subséquente d'un photon amène à la formation d'un trion localisé sur les charges piégées. / In this dissertation we report on original experimental investigations of the photophysical properties of individual single-walled carbon nanotubes (SWCNTs) at the single molecule level. We focused on two problems: the effect of water-filling nanotubes on their photoluminescence (PL) properties and generation of multi-excitonic complexes in regimes of strong laser excitation. To do so we used a combination of microscopy, PL spectroscopy and PL time decay measurements. Our results show, for different nanotube diameters, a red-shift of the PL emission energy for water-filled nanotubes. Furthermore, biexponential PL time decay behaviors are obtained for individual water-filled and empty (6,4) SWCNTs. The characteristic decay times for both species of nanotubes are distinct, with a reduction of the short component in water-filled SWCNTs. These results are explained by an increase of the dielectric constant for water-filled nanotubes. Our most consequential experimental result was the observation of all optical trion generation in undoped nanotubes. Trion emission appears in the PL spectrum as a red-shifted sideband. Based on our experimental observations, we propose a trion generation model according to which, at a multiexcitonic regime, exciton-exciton annihilation interactions create charge carriers that are transiently trapped at electrostatic potential fluctuations induced by the inhomogeneities of the environment. Subsequent photon absorption leads to trion formation localized at the trapped carriers.

The modelling of single-molecule experiments is of vital interest to gain new insights into processes which were hitherto not accessible by measurements performed on bulk systems. In the first part of this thesis, the kinetics of a triple-branch DNA molecule with four conformational states is investigated by employing pulling experiments with optical tweezers and theoretical modelling. Probability distributions of first rupture forces, which are calculated by applying transition rate theory to a free energy model, show good agreement with experimental findings. Permanently frayed molecules could be identified by analysing the number of opening base pairs in force jumps. In the second part of the thesis, DNA hairpin molecules with periodic base sequences are studied. Their unfolding kinetics allows an analytical treatment, because they exhibit a regular coarse-grained free energy landscape as a function of the number of opened base pairs. A procedure is developed for determining all relevant parameters of the landscape, which relies on probabilities that can be easily sampled from the unfolding trajectories. By means of Monte Carlo simulations it is shown that already 300 trajectories, as typically measured in single-molecule experiments, provide faithful results for the energetic parameters. The approach in particular opens a new access to improve loop contributions in the free energy landscape. In the third part of the thesis, a simulation method is developed for modelling the unfolding kinetics of DNA molecules with arbitrary base sequences. The method is validated against experimental data for five DNA hairpin molecules with different length of the end-loop. Applications of the method enable one, among others, to improve the parameter determination in functional forms suggested for the tail behaviour of work distributions. Such work distributions enter detailed and integral fluctuation theorems, which are useful for estimating free energy differences between folded and unfolded states from nonequilibrium measurements.

No description available.