Suchergebnisse

Research on biosensors is growing in relevance, taking benefit from groundbreaking knowledge that allows for new biosensing strategies. Electrochemical biosensors can benefit from research on semiconducting materials for energy applications. This research seeks the optimization of the semiconductor-electrode interfaces including light-harvesting materials, among other improvements. Once that knowledge is acquired, it can be implemented with biological recognition elements, which are able to transfer a chemical signal to the photoelectrochemical system, yielding photo-biosensors. This has been a matter of research as it allows both a superior suppression of background electrochemical signals and the switching ON and OFF upon illumination. Effective electrode-semiconductor interfaces and their coupling with biorecognition units are reviewed in this work. ; European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No. 713366. G. Luna-López wishes to thank to "Comunidad de Madrid" and European Structural Funds for their financial support to FotoArt-CM project (S2018/NMT-4367). M. Pita thanks the Retos MCIU/AEI/FEDER, EU for funding project RTI2018-095090-B-I00.

The successive steps of laccase immobilization on chemically modified Au electrodes were monitored using ATR-SEIRAS and in situ STM. Successful covalent immobilization of the enzyme on Au electrodes modified by a mixed aminophenyl-mercaptohexanol adlayer and on Au electrodes modified by a 4-aminothiophenyl SAM via a Schiff base reaction followed by the formation of amide bonds is revealed by the emergence of the corresponding bands in the ATR-SEIRA spectra, and an enzyme coverage on aminophenyl-mercaptohexanol- modified Au electrodes of about (7.27 ± 1.93) × 10 11 laccase units per cm 2 was calculated from STM images. The small differences between the ATR-SEIRA spectra of the enzyme immobilized on aminophenyl-mercaptohexanol-modified Au electrodes and the ATR-SEIRA spectra of the enzyme immobilized on 4-aminothiophenyl-modified Au electrodes are attributed to a different orientation of the immobilized enzyme due to the presence on the surface of aminophenyl-mercaptohexanol-modified Au electrodes of OH functional groups that favor an orientation of laccase with the Cu T1 center of the enzyme facing the electrode surface, thus, allowing a high activity for direct electrocatalysis of the ORR at low overpotentials. © 2012 American Chemical Society. ; This work has been funded by the DGI (Ministerio de Ciencia e Innovación), under Projects CTQ2009-07017 and CTQ2009-12649, and by the European Union (FP7 Project "3D-Nanobiodevice", NMP4-SL-2009-229255). C.V.-D. acknowledges a JAE-Doc fellowship from CSIC, and M.P. acknowledges the Ramon y Cajal 2009 program from the Spanish MICINN. ; Peer Reviewed

[EN] NADH is a key biomolecule involved in many biocatalytic processes as cofactor and its quantification can be correlated to specific enzymatic activity. Many effort s have been t aken to obt ain clean electrochemical signals related to NADH presence and lower its redox overpotential to avoid interferences. Suppression of background and secondary signals can be achieved by including a switchable electroactive surface, for instance, by using semiconductors able to harvest light energy and drive the excited electrons only when irradiated. Here we present the combination of a n-type Si semiconductor with fibers made of carbon nanotubes as electroactive surface for NADH quantification at low potentials only upon irradiation. The resulting photoelectrode responded linearly to NADH concentrations from 50 μM to 1.6 mM with high sensitivity (54 μA cm −2 mM −1 ). This system may serve as a biosensing platform for detection and quantification of dehydrogenases' activity. ; European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No. 713366 . A.D.L., M.R and J.J.V. thank the "Comunidad de Madrid "for its support to the FotoArt-CM project ( S2018/NMT-4367 ) through the Program of R&D activities between research groups in Technologies 2018, co- financed by European Structural Funds. J.J.V. is grateful for generous financial support provided by the European Union Horizon 2020 Program under grant agreement 678565 (ERC-STEM) and by the MINECO ( RyC-2014-15115 HYNANOSC RTI2018-099504-A-C22 ; Peer reviewed

High surface area graphene electrodes were prepared by simultaneous electrodeposition and electroreduction of graphene oxide. The electrodeposition process was optimized in terms of pH and conductivity of the solution and the obtained graphene electrodes were characterized by X-ray photoelectron spectroscopy, Fourier transform infrared spectroscopy, scanning electron microscopy and electrochemical methods (cyclic voltammetry and impedance spectroscopy). Electrodeposited electrodes were further functionalized to carry out covalent immobilization of two oxygen-reducing multicopper oxidases: laccase and bilirubin oxidase. The enzymatic electrodes were tested as direct electron transfer based biocathodes and catalytic currents as high as 1 mA/cm2 were obtained. Finally, the mechanism of the enzymatic oxygen reduction reaction was studied for both enzymes calculating the Tafel slopes and transfer coefficients. ; This work has received funding from the European Union's Seventh Framework Programme under grant agreement BIOENERGY, FP7-PEOPLE-2013-ITN-607793. ; Peer Reviewed

High-redox potential laccases are powerful biocatalysts with a wide range of applications in biotechnology. We have converted a thermostable laccase from a white-rot fungus into a blood tolerant laccase. Adapting the fitness of this laccase to the specific composition of human blood (above neutral pH, high chloride concentration) required several generations of directed evolution in a surrogate complex blood medium. Our evolved laccase was tested in both human plasma and blood, displaying catalytic activity while retaining a high redox potential at the T1 copper site. Mutations introduced in the second coordination sphere of the T1 site shifted the pH activity profile and drastically reduced the inhibitory effect of chloride. This proof of concept that laccases can be adapted to function in extreme conditions opens an array of opportunities for implantable nanobiodevices, chemical syntheses, and detoxification. ; This study was based upon work funded by EU Projects (NMP4-SL-2009-229255-3D-Nanobiodevice, FP7-KBBE-2010-4-26537-Peroxicats and COST Action CM0701) and a project from the Spanish Government (BIO2010-19697). D.M.M. was supported by a JAE grant and D.G.P. was supported by a Peroxicats contract. M.P. received support from the 2009 Ramón y Cajal programme of the Spanish MINECO. ; Peer reviewed

Enzyme engineering has allowed not only the de novo creation of active sites catalysing known biological reactions with rates close to diffusion limits, but also the generation of abiological sites performing new-to-nature reactions. However, the catalytic advantages of engineering multiple active sites into a single protein scaffold are yet to be established. Here, we report on proteins with two active sites of biological and/or abiological origin, for improved natural and non-natural catalysis. The approach increased the catalytic properties, such as enzyme efficiency, substrate scope, stereoselectivity and optimal temperature window, of an esterase containing two biological sites. Then, one of the active sites was metamorphosed into a metal-complex chemocatalytic site for oxidation and Friedel–Crafts alkylation reactions, facilitating synergistic chemo- and biocatalysis in a single protein. The transformations of 1-naphthyl acetate into 1,4-naphthoquinone (conversion approx. 100%) and vinyl crotonate and benzene into 3-phenylbutyric acid (≥83%; e.e. >99.9%) were achieved in one pot with this artificial multifunctional metalloenzyme. ; This work was funded by grant 'INMARE' from the European Union's Horizon 2020 (grant agreement no. 634486), grants PCIN-2017-078 (within the Marine Biotechnology ERA-NET), CTQ2016-79138-R, BIO2016-76601-C3-1-R, BIO2016-76601-C3-3-R, BIO2017-85522-R, RTI2018-095166-B-I00 and RTI2018-095090-B-100 from the Ministerio de Economía y Competitividad, the Ministerio de Ciencia, Innovación y Universidades (MCIU), the Agencia Estatal de Investigación (AEI), the Fondo Europeo de Desarrollo Regional (FEDER) and the European Union (EU). P.N.G. and R.B. acknowledge the support of the UK Biotechnology and Biological Sciences Research Council (BBSRC; grant No. BB/M029085/1) and the Centre of Environmental Biotechnology Project and the Supercomputing Wales project, which are partly funded by the European Regional Development Fund (ERDF) through the Welsh Government. The authors gratefully acknowledge the financial support provided by the ERDF. C.C. thanks the Ministerio de Economía y Competitividad and FEDER for a Ph.D. fellowship (Grant BES-2015-073829). J.L.G.-A. thanks the support of the Spanish Ministry of Education, Culture and Sport through the National Program FPU (FPU17/00044). I.C.-R. thanks the Regional Government of Madrid for a fellowship (PEJ_BIO_AI_1201). The authors would like to acknowledge S. Ciordia and M. C. Mena for MALDI-TOF/TOF analysis. We thank the staff of both the European Synchrotron Radiation Facility (ESRF, Grenoble, France), for providing access and technical assistance at beamline ID30A-1/MASSIf-1, and the Synchrotron Radiation Source at Alba (Barcelona, Spain), for assistance at BL13-XALOC beamline. The authors would also like to acknowledge M. J. Vicente and M. A. Pascual at the Servicio Interdepartamental de Investigación (SIDI) of the Autonomous University of Madrid for the ESI-MS analyses.