title: Proton-coupled electron transfer of flavodoxin immobilized on nanostructured tin dioxide electrodes: thermodynamics versus kinetics control of protein redox function.
abstract: In this paper, we report a spectroelectrochemical investigation of proton-coupled electron transfer in flavodoxin D. vulgaris Hildenborough (Fld). Poly-L-lysine is used to promote the binding of Fld to the nanocrystalline, mesoporous SnO(2) electrodes. Two reversible redox couples of the immobilized Fld are observed electrochemically and are assigned by spectroelectrochemistry to the quinone/semiquinone and semiquinone/hydroquinone couples of the protein's flavin mononucleotide (FMN) redox cofactor. Comparison with control data for free FMN indicates no contamination of the Fld data by dissociated FMN. The quinone/semiquinone and semiquinone/hydroquinone midpoint potentials (E(q/sq) and E(sq/hq)) at pH 7 were determined to be -340 and -585 mV vs Ag/AgCl, in good agreement with the literature. E(q/sq) exhibited a pH dependence of 51 mV/pH. The kinetics of these redox couples were studied using cyclic voltammetry, cyclic voltabsorptometry, and chronoabsorptometry. The semiquinone/quinone reoxidation is found to exhibit slow, potential-independent but pH-sensitive kinetics with a reoxidation rate constant varying from 1.56 s(-)(1) at pH 10 to 0.0074 s(-)(1) at pH 5. The slow kinetics are discussed in terms of a simple kinetics model and are assigned to the reoxidation process being rate limited by semiquinone deprotonation. It is proposed that this slow deprotonation step has the physiological benefit of preventing the undesirable loss of reducing equivalents which results from semiquinone oxidation to quinone.
authors.affiliation: Department of Chemistry, Imperial College London, South Kensington, London SW7 2AZ, UK.
authors.forename: Yeni
authors.initials: Y
authors.lastname: Astuti
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Topoglidis
authors.forename: Paul B
authors.initials: PB
authors.lastname: Briscoe
authors.forename: Andrea
authors.initials: A
authors.lastname: Fantuzzi
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Gilardi
authors.forename: James R
authors.initials: JR
authors.lastname: Durrant
mesh_terms: D003460:Crystallization; D004566:Electrodes; D004579:Electron Transport; D005486:Flavin Mononucleotide / Q000737:chemistry / Q000378:metabolism; D005182:Flavin-Adenine Dinucleotide / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D005418:Flavodoxin / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006873:Hydroquinones / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008832:Microchemistry; D010084:Oxidation-Reduction; D011107:Polylysine / Q000737:chemistry; D011522:Protons; D011809:Quinones / Q000737:chemistry / Q000378:metabolism; D013816:Thermodynamics; D017971:Tin Compounds / Q000737:chemistry*
chemical_list: D005418:Flavodoxin; D006873:Hydroquinones; D011522:Protons; D011809:Quinones; D017971:Tin Compounds; D005182:Flavin-Adenine Dinucleotide; D011107:Polylysine; C029276:flavin semiquinone; D005486:Flavin Mononucleotide; C045358:stannic oxide