Specific and reversible immobilization of proteins tagged to the affinity polypeptide C-LytA on functionalized graphite electrodes

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Título: Specific and reversible immobilization of proteins tagged to the affinity polypeptide C-LytA on functionalized graphite electrodes
Autor/es: Bello Gil, Daniel | Maestro, Beatriz | Fonseca, Jennifer | Feliu, Juan M. | Climent, Victor | Sanz, Jesús M.
Grupo/s de investigación o GITE: Electroquímica de Superficies
Centro, Departamento o Servicio: Universidad de Alicante. Departamento de Química Física | Universidad de Alicante. Instituto Universitario de Electroquímica
Palabras clave: Specific and reversible immobilization | Proteins | C-LytA | Graphite electrodes
Área/s de conocimiento: Química Física
Fecha de publicación: 31-ene-2014
Editor: Public Library of Science (PLoS)
Cita bibliográfica: Bello-Gil D, Maestro B, Fonseca J, Feliu JM, Climent V, et al. (2014) Specific and Reversible Immobilization of Proteins Tagged to the Affinity Polypeptide C-LytA on Functionalized Graphite Electrodes. PLoS ONE 9(1): e87995. doi:10.1371/journal.pone.0087995
Resumen: We have developed a general method for the specific and reversible immobilization of proteins fused to the choline-binding module C-LytA on functionalized graphite electrodes. Graphite electrode surfaces were modified by diazonium chemistry to introduce carboxylic groups that were subsequently used to anchor mixed self-assembled monolayers consisting of N,N-diethylethylenediamine groups, acting as choline analogs, and ethanolamine groups as spacers. The ability of the prepared electrodes to specifically bind C-LytA-tagged recombinant proteins was tested with a C-LytA-β-galactosidase fusion protein. The binding, activity and stability of the immobilized protein was evaluated by electrochemically monitoring the formation of an electroactive product in the enzymatic hydrolysis of the synthetic substrate 4-aminophenyl β-D-galactopyranoside. The hybrid protein was immobilized in an specific and reversible way, while retaining the catalytic activity. Moreover, these functionalized electrodes were shown to be highly stable and reusable. The method developed here can be envisaged as a general, immobilization procedure on the protein biosensor field.
Patrocinador/es: This work was supported by grants BFU2010–17824, CTQ2008-04492-E and CTQ2010-18570 (Spanish Ministerio de Economia y Competitividad) and FPA-2010-055 (Valentian Government, Spain).
URI: http://hdl.handle.net/10045/35464
ISSN: 1932-6203
DOI: 10.1371/journal.pone.0087995
Idioma: eng
Tipo: info:eu-repo/semantics/article
Derechos: © 2014 Bello-Gil et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Revisión científica: si
Versión del editor: http://dx.doi.org/10.1371/journal.pone.0087995
Aparece en las colecciones:INV - EQSUP - Artículos de Revistas

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