Fully integrated biocatalytic electrodes based on bioaffinity interactions

Eugenii Katz, Vered Heleg-Shabtai, Amos Bardea, Itamar Willner, Harald K. Rau, Wolfgang Haehnel

Research output: Contribution to journalConference articlepeer-review

62 Scopus citations

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 Co(II)- protoporphyrin IX reconstituted myoglobin (Co(II)-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 Co(II)-Mb show catalytic activities for NO3- reduction and acetylenedicarboxylic acid hydrogenation, respectively. Two Fe(III)- protoporphyrin IX units are reconstituted into a four α-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 Fe(III)-protoporphyrin IX reconstituted de novo protein monolayer and NR yields an integrated, electrically contacted enzyme electrode that stimulates the bioelectrocatalyzed reduction of nitrate.

Original languageEnglish
Pages (from-to)741-756
Number of pages16
JournalBiosensors and Bioelectronics
Volume13
Issue number7-8
DOIs
StatePublished - 1 Oct 1998
Externally publishedYes
EventProceedings of the 1998 5th World Congress on Biosensors - Berlin, Ger
Duration: 3 Jun 19985 Jun 1998

Keywords

  • Bioelectronics
  • Biosensor
  • De novo proteins
  • Electrical contact of enzymes
  • Enzyme monolayers
  • Integrated enzyme electrodes
  • Microperoxidase-11
  • Myoglobin
  • NAD-dependent enzymes
  • Nitrate reductase
  • PQQ
  • Pyrroloquinoline quinone
  • Reconstituted enzymes

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