Nanomechanics: A new approach for studying the mechanical properties of materials

Mitjançant l'espectroscòpia de forces atòmiques s'ha estudiat la resposta nanomecànica a la nanoindentació de la superfície més estable d'un material trencadís FCC, com és ara el MgO (100). L'expulsió del material en forma de capes demostra que la fallida trencadissa implica, de fet, l'inici de la deformació plàstica o estrès crític, i que la deformació plàstica posterior consisteix en una sèrie d'esdeveniments discrets. Es pot determinar amb precisió el mòdul de Young, E, a partir de la regió de deformació elàstica mitjançant una mecànica senzilla, atesa l'absència de dislocacions induïdes per la nanoindentació. Amb aquesta finalitat s'ha desenvolupat un nou model fisicomatemàtic, que té en compte les interaccions laterals. El valor de l'estrès crític de fricció també s'ha calculat i comentat. Com a conseqüència d'aquesta expulsió en capes, també s'ha estudiat la resposta nanomecànica de superfícies de capes primes (gruix & 1 µm) de molècules orgàniques altament orientades, ja que es tracta de materials en capes amb interaccions de tipus Van der Waals. També en aquests materials la superfície es deforma plàsticament i presenta discontinuïtats discretes en les corbes d'indentació, associades ara a les capes moleculars expulsades per l'indentador. En el cas del metall quasiunidimensional tetratiofulvalè tetracianoquinodimetà (TTFTCNQ), el valor del mòdul de Young, E & 20 GPa, coincideix amb l'obtingut per altres mètodes. En el cas de la fase ! del radical p-nitrofenil nitronil nitròxid (p-NPNN) no es disposa d'informació per a monocristalls, i el valor obtingut per a les capes primes és de E & 2 GPa.Atomic force spectroscopy was used to study the nanomechanical response to nanoindentations on the most stable face (100) of FCC brittle materials such as MgO and alkali halides. The layered expulsion of material demonstrates that brittle failure results from the critical stress brought on by plastic deformation and that plastic deformation consists of a series of discrete events. Due to the absence of indentation- induced dislocations, Young?s modulus E can be correctly estimated from the elastic deformation region using simple mechanics. A new model is developed taking into account lateral interactions. Critical shear stress is also evaluated and discussed. As a result of the layered expulsion we also studied the nanomechanical response of surfaces of highly-oriented molecular organic thin films (ca. 1 µm thickness) because these are Van der Waals layered materials. The surfaces were again found to deform plastically and there were discrete discontinuities in the indentation curves, representing the molecular layers being expelled by the penetrating tip. Here, the Hertz model is quite good at revealing the role of lateral interactions in the indentation process. For the quasi-one-dimensional metal tetrathiafulvalene tetracyanoquinodimethane (TTF-TCNQ) the value of Young?s modulus, E & 20 GPa, coincides with that obtained by other bulk methods. For the !-phase of the p-nitrophenyl nitronyl nitroxide (p-NPNN) radical, no information is available for single crystals and the estimated value obtained for the film is E & 2 GPa

Long distance electron transfer through the aqueous solution between redox partner proteins

Despite the importance of electron transfer between redox proteins in photosynthesis and respiration, the inter-protein electron transfer rate between redox partner proteins has never been measured as a function of their separation in aqueous solution. Here, we use electrochemical tunneling spectroscopy to show that the current between two protein partners decays along more than 10 nm in the solution. Molecular dynamics simulations reveal a reduced ionic density and extended electric field in the volume confined between the proteins. The distance-decay factor and the calculated local barrier for electron transfer are regulated by the electrochemical potential applied to the proteins. Redox partners could use electrochemically gated, long distance electron transfer through the solution in order to conciliate high specificity with weak binding, thus keeping high turnover rates in the crowded environment of cells.España, MINECO BFU2015-71017-P/España, Gobierno de Andalucía (BIO198)Comisión de Universidades e Investigación del Departamento de Innovación, Universidades y Empresa de la Generalitat de Catalunya.(2017 SGR 1442, 2014 SGR 1442, and 2017 SGR 1189

Direct Measurement of the Nanomechanical Stability of a Redox Protein Active Site and Its Dependence upon Metal Binding

The structural basis of the low reorganization energy of cupredoxins has long been debated. These proteins reconcile a conformationally heterogeneous and exposed metal-chelating site with the highly rigid copper center required for efficient electron transfer. Here we combine single-molecule mechanical unfolding experiments with statistical analysis and computer simulations to show that the metal-binding region of apo-azurin is mechanically flexible and that high mechanical stability is imparted by copper binding. The unfolding pathway of the metal site depends on the pulling residue and suggests that partial unfolding of the metal binding site could be facilitated by the physical interaction with certain regions of the redox protein.We are grateful to A. Donaire and I. Díez-Pérez for discussions, and to the Catalan government (grant 2014SGR-1251), the Spanish government (grant CTQ2013-43892R) and the European Research Council (PELE ERC-2009-Adg 25027) for financial support.Peer ReviewedPostprint (author's final draft

Fast Photoswitchable Molecular Prosthetics Control Neuronal Activity in the Cochlea

Altres ajuts: CERCA Programme/Generalitat de Catalunya, Fundaluce and "la Caixa" foundations (ID 100010434, agreement LCF/PR/HR19/52160010)Artificial control of neuronal activity enables the study of neural circuits and restoration of neural functions. Direct, rapid, and sustained photocontrol of intact neurons could overcome the limitations of established electrical stimulation such as poor selectivity. We have developed fast photoswitchable ligands of glutamate receptors (GluRs) to enable neuronal control in the auditory system. The new photoswitchable ligands induced photocurrents in untransfected neurons upon covalently tethering to endogenous GluRs and activating them reversibly with visible light pulses of a few milliseconds. As a proof of concept of these molecular prostheses, we applied them to the ultrafast synapses of auditory neurons of the cochlea that encode sound and provide auditory input to the brain. This drug-based method afforded the optical stimulation of auditory neurons of adult gerbils at hundreds of hertz without genetic manipulation that would be required for their optogenetic control. This indicates that the new photoswitchable ligands are also applicable to the spatiotemporal control of fast spiking interneurons in the brain

Fast photochronoamperometry of photosynthetic complexes for biosensors and electron transport studies

Photosynthetic reactions in plants, algae, and cyanobacteria are driven by photosystem I and photosystem II complexes, which specifically reduce or oxidize partner redox biomolecules. Photosynthetic complexes can also bind synthetic organic molecules, which inhibit their photoactivity and can be used both to study the electron transport chain and as herbicides and algicides. Thus, their development, characterization, and sensing bears fundamental and applied interest. Substantial efforts have been devoted to developing photosensors based on photosystem II to detect compounds that bind to the plastoquinone sites of this complex. In comparison, photosystem I based sensors have received less attention and could be used to identify novel substances displaying phytotoxic effects, including those obtained from natural product extracts. We have developed a robust procedure to functionalize gold electrodes with photo- and redox-active photosystem I complexes based on transparent gold and a thiolate self-assembled monolayer, and we have obtained reproducible electrochemical photoresponses. Chronoamperometric recordings have allowed us to measure photocurrents in the presence of the viologen derivative paraquat at concentrations below 100 nM under lock-in operation and a sensor dynamic range spanning six orders of magnitude up to 100 mM. We have modeled their time course to identify the main electrochemical processes and limiting steps in the electron transport chain. Our results allow us to isolate the contributions from photosystem I and the redox mediator, and evaluate photocurrent features (spectral and power dependence, fast transient kinetics) that could be used as a sensing signal to detect other inhibitors and modulators of photosystem I activity.Peer ReviewedPostprint (author's final draft

Positional isomers of bispyridine benzene derivatives induce efficacy changes on mGlu5 negative allosteric modulation

Modulation of metabotropic glutamate receptor 5 (mGlu5) with partial allosteric antagonists has received increased interest due to their favourable in vivo activity profiles compared to the unfavourable side-effects of full inverse agonists. Here we report on a series of bispyridine benzene derivatives with a functional molecular switch affecting antagonistic efficacy, shifting from inverse agonism to partial antagonism with only a single change in the substitution pattern of the benzene ring. These efficacy changes are explained through computational docking, revealing two different receptor conformations of different energetic stability and different positional isomer binding preferences

In vivo photocontrol of orexin receptors with a nanomolar light-regulated analogue of orexin-B

Gorostiza, Pa

Shining light on an mGlu5 photoswitchable NAM: A theoretical perspective

Metabotropic glutamate receptors (mGluRs) are important drug targets because of their involvement in several neurological diseases. Among mGluRs, mGlu5 is a particularly high-profile target because its positive or negative allosteric modulation can potentially treat schizophrenia or anxiety and chronic pain, respectively. Here, we computationally and experimentally probe the functional binding of a novel photoswitchable mGlu5 NAM, termed alloswitch-1, which loses its NAM functionality under violet light. We show alloswitch-1 binds deep in the allosteric pocket in a similar fashion to mavoglurant, the co-crystallized NAM in the mGlu5 transmembrane domain crystal structure. Alloswitch-1, like NAM 2-Methyl-6-(phenylethynyl)pyridine (MPEP), is significantly affected by P655M mutation deep in the allosteric pocket, eradicating its functionality. In MD simulations, we show alloswitch-1 and MPEP stabilize the co-crystallized water molecule located at the bottom of the allosteric site that is seemingly characteristic of the inactive receptor state. Furthermore, both NAMs form H-bonds with S809 on helix 7, which may constitute an important stabilizing interaction for NAM-induced mGlu5 inactivation. Alloswitch-1, through isomerization of its amide group from trans to cis is able to form an additional interaction with N747 on helix 5. This may be an important interaction for amide-containing mGlu5 NAMs, helping to stabilize their binding in a potentially unusual cis-amide state. Simulated conformational switching of alloswitch-1 in silico suggests photoisomerization of its azo group from trans to cis may be possible within the allosteric pocket. However, photoexcited alloswitch-1 binds in an unstable fashion, breaking H-bonds with the protein and destabilizing the co-crystallized water molecule. This suggests photoswitching may have destabilizing effects on mGlu5 binding and functionality

Large-Scale Dendrimer-Based Uneven Nanopatterns for the Study of Local RGD Density Effects on Cell Adhesion

Cell adhesion processes are governed by the nanoscale arrangement of the extracellular matrix (ECM), being more affected by local rather than global concentrations of cell adhesive ligands. In many cell-based studies, grafting of dendrimers on surfaces has shown the benefits of the local increase in concentration provided by the dendritic configuration, although the lack of any reported surface characterization has limited any direct correlation between dendrimer disposition and cell response. In order to establish a proper correlation, some control over dendrimer surface deposition is desirable. Here, dendrimer nanopatterning has been employed to address arginine–glycine–aspartic acid (RGD) density effects on cell adhesion. Nanopatterned surfaces were fully characterized by atomic force microscopy (AFM), scanning tunneling microscopy (STM) and X-ray photoelectron spectroscopy (XPS), showing that tunable distributions of cell adhesive ligands on the surface are obtained as a function of the initial dendrimer bulk concentration. Cell experiments showed a clear correlation with dendrimer surface layout: Substrates presenting regions of high local ligand density resulted in a higher percentage of adhered cells and a higher degree of maturation of focal adhesions (FAs). Therefore, dendrimer nanopatterning is presented as a suitable and controlled approach to address the effect of local ligand density on cell response. Moreover, due to the easy modification of dendrimer peripheral groups, dendrimer nanopatterning can be further extended to other ECM ligands having density effects on cells