title: Fully integrated biocatalytic electrodes based on bioaffinity interactions.
abstract: Integrated bioelectrocatalytically active electrodes are assembled by the deposition of enzymes onto respective electrically contacted affinity matrices and further cross-linking of the enzyme monolayers. A catalyst-NAD(+)-dyad for the binding of the NAD(+)-dependent enzymes and cytochrome-like molecules for the binding of the heme-protein-dependent enzymes are used to construct integrated electrically contacted biocatalytic systems. NAD(+)-dependent lactate dehydrogenase (LDH) is assembled onto a pyrroloquinoline quinone-NAD+ monolayer. The redox-active monolayer is organized via covalent attachment of pyrroloquinoline quinone (PQQ) to a cystamine monolayer associated with a Au-electrode, followed by covalent linkage of N6-(2-aminoethyl)-NAD+ to the monolayer. The interface modified with the PQQ-NAD(+)-dyad provides temporary affinity binding for LDH and allows cross-linking of the enzyme monolayer. The cross-linked LDH is bioelectrocatalytically active towards oxidation of lactate. The bioelectrocatalyzed process involves the PQQ-mediated oxidation of the immobilized NADH. Integrated, electrically contacted bioelectrodes are produced by the affinity binding and further cross-linking of nitrate reductase (NR) (cytochrome-dependent, E.C. 1.9.6.1 from E. coli) or CoII-protoporphyrin IX reconstituted myoglobin (CoII-Mb) atop the microperoxidase-11 (MP-11) monolayer associated with a Au-electrode. The MP-11 monolayer provides an affinity interface for the temporary binding of the enzymes, that allows the cross-linkage of the enzyme molecules. The MP-11 assembly acts as electron transfer mediator for the reduction of the secondary enzyme layer. The integrated bioelectrodes consisting of NR and CoII-Mb show catalytic activities for NO3- reduction and acetylene-dicarboxylic acid hydrogenation, respectively. Two FeIII-protoporphyrin IX units are reconstituted into a four alpha-helix bundle de novo protein assembled as a monolayer on a Au-electrode. Vectorial electron transfer proceeds in the synthetic heme-protein monolayer. Cross-linking of an affinity complex generated between the FeIII-protoporphyrin IX reconstituted de novo protein monolayer and NR yields an integrated, electrically contacted enzyme electrode that stimulates the bioelectrocatalyzed reduction of nitrate.
authors.affiliation: Institute of Chemistry, Hebrew University of Jerusalem, Israel. ekatz@vms.huji.ac.il
authors.forename: E
authors.initials: E
authors.lastname: Katz
authors.forename: V
authors.initials: V
authors.lastname: Heleg-Shabtai
authors.forename: A
authors.initials: A
authors.lastname: Bardea
authors.forename: I
authors.initials: I
authors.lastname: Willner
authors.forename: H K
authors.initials: HK
authors.lastname: Rau
authors.forename: W
authors.initials: W
authors.lastname: Haehnel
mesh_terms: D000818:Animals; D015374:Biosensing Techniques; D002384:Catalysis; D004798:Enzymes / Q000032:analysis* / Q000737:chemistry; D006801:Humans; D010084:Oxidation-Reduction; D055672:Static Electricity
chemical_list: D004798:Enzymes