title: The two-step electrochemical oxidation of alcohols using a novel recombinant PQQ alcohol dehydrogenase as a catalyst for a bioanode.
abstract: A bioanode has been developed based on the oxidation of ethanol by the recombinant pyrroloquinoline quinone (PQQ) dependent alcohol dehydrogenase from Pseudomonas putidaKT2440 heterologously expressed in Pichia pastoris. The apo form of the recombinant protein (PpADH) was purified and displayed catalytic activity for binding PQQ in the presence of Ca(2+). PpADH exhibited broad substrate specificity towards various alcohols and aldehydes. The Km values for the aldehydes of PpADH were increased compared to those for the alcohols, whereas the kcat values were unaltered. For instance, the Km values at T=298.15K (25 °C) for ethanol and acetaldehyde were 0.21 (± 0.02)mM and 5.8 (± 0.60)mM, respectively. The kcat values for ethanol and acetaldehyde were 24.8 (± 1.2) s(-1) and 31.1 (± 1.2) s(-1), respectively. The aminoferrocene was used as an electron transfer mediator between PpADH and the electrode during electrochemical experiments. The catalytic currents for the oxidation of alcohol and acetaldehyde by PpADH were also observed in this system. The electric charge for the oxidation of ethanol (Q = 2.09 × 10(-3) · C) was increased two-fold compared to that for the oxidation of acetaldehyde (Q = 0.95 × 10(-3) · C), as determined by chronoamperometric measurements. Thus, we have electrochemically demonstrated the two-step oxidation of ethanol to acetate using only PpADH.
authors.affiliation: Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan.
authors.forename: Kouta
authors.initials: K
authors.lastname: Takeda
authors.forename: Hirotoshi
authors.initials: H
authors.lastname: Matsumura
authors.forename: Takuya
authors.initials: T
authors.lastname: Ishida
authors.forename: Masahiro
authors.initials: M
authors.lastname: Samejima
authors.forename: Kiyohiko
authors.initials: K
authors.lastname: Igarashi
authors.forename: Nobuhumi
authors.initials: N
authors.lastname: Nakamura
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Ohno
mesh_terms: D000438:Alcohols / Q000737:chemistry* / Q000378:metabolism; D002237:Carbohydrate Dehydrogenases / Q000737:chemistry* / Q000235:genetics; D002384:Catalysis; D004563:Electrochemistry; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D010084:Oxidation-Reduction; D045542:PQQ Cofactor / Q000737:chemistry*; D010843:Pichia / Q000235:genetics; D016958:Pseudomonas putida / Q000201:enzymology
chemical_list: D000438:Alcohols; D045542:PQQ Cofactor; D002237:Carbohydrate Dehydrogenases; C082601:aldose dehydrogenase (pyrroloquinoline-quinone)
keywords: Electrocatalysis; Enzymatic biofuel cell; Mediated electron transfer; Pyrroloquinoline quinone; Quinoprotein