title: Investigation of protein FTT1103 electroactivity using carbon and mercury electrodes. Surface-inhibition approach for disulfide oxidoreductases using silver amalgam powder.
abstract: Recently, it was shown that electrochemical methods can be used for analysis of poorly water-soluble proteins and for study of their structural changes and intermolecular (protein-ligand) interactions. In this study, we focused on complex electrochemical investigation of recombinant protein FTT1103, a disulfide oxidoreductase with structural similarity to well described DsbA proteins. This thioredoxin-like periplasmic lipoprotein plays an important role in virulence of bacteria Francisella tularensis. For electrochemical analyses, adsorptive transfer (ex situ) square-wave voltammetry with pyrolytic graphite electrode, and alternating-current voltammetry and constant-current chronopotentiometric stripping analysis with mercury electrodes, including silver solid amalgam electrode (AgSAE) were used. AgSAE was used in poorly water-soluble protein analysis for the first time. In addition to basic redox, electrocatalytic and adsorption/desorption characterization of FTT1103, electrochemical methods were also used for sensitive determination of the protein at nanomolar level and study of its interaction with surface of AgSA microparticles. Proposed electrochemical protocol and AgSA surface-inhibition approach presented here could be used in future for biochemical studies focused on proteins associated with membranes as well as on those with disulfide oxidoreductase activity.
authors.affiliation: Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacký University, Hnevotinska 3, Olomouc 775 15, Czech Republic.
authors.forename: Renata
authors.initials: R
authors.lastname: Večerková
authors.affiliation: Regional Centre for Applied Molecular Oncology, Masaryk Memorial Cancer Institute, Zluty kopec 7, Brno 656 53, Czech Republic.
authors.forename: Lenka
authors.initials: L
authors.lastname: Hernychová
authors.affiliation: Regional Centre for Applied Molecular Oncology, Masaryk Memorial Cancer Institute, Zluty kopec 7, Brno 656 53, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Dobeš
authors.affiliation: Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacký University, Hnevotinska 3, Olomouc 775 15, Czech Republic.
authors.forename: Jiří
authors.initials: J
authors.lastname: Vrba
authors.affiliation: J. Heyrovský Institute of Physical Chemistry of AS CR, v.v.i., Department of Biomimetic Electrochemistry, Dolejskova 3, Prague 182 23, Czech Republic.
authors.forename: Bohdan
authors.initials: B
authors.lastname: Josypčuk
authors.affiliation: Regional Centre for Applied Molecular Oncology, Masaryk Memorial Cancer Institute, Zluty kopec 7, Brno 656 53, Czech Republic.
authors.forename: Martin
authors.initials: M
authors.lastname: Bartošík
authors.affiliation: Department of Medical Chemistry and Biochemistry, Faculty of Medicine and Dentistry, Palacký University, Hnevotinska 3, Olomouc 775 15, Czech Republic. Electronic address: jan.vacek@upol.cz.
authors.forename: Jan
authors.initials: J
authors.lastname: Vacek
mesh_terms: D000327:Adsorption; D002244:Carbon / Q000737:chemistry*; D055664:Electrochemical Techniques; D004566:Electrodes; D008628:Mercury / Q000737:chemistry*; D008958:Models, Molecular; D010316:Particle Size; D011208:Powders / Q000737:chemistry; D011490:Protein Disulfide Reductase (Glutathione) / Q000032:analysis* / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000378:metabolism; D012834:Silver / Q000737:chemistry* / Q000494:pharmacology; D013499:Surface Properties
chemical_list: D011208:Powders; D011994:Recombinant Proteins; D012834:Silver; D002244:Carbon; D011490:Protein Disulfide Reductase (Glutathione); D008628:Mercury
keywords: Disulfide bond forming protein; Electrochemical sensing; Membrane proteins; Oxidoreductase; Surface inhibition