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Monolayers of photosystem II on gold electrodes with enhanced sensor response—effect of porosity and protein layer arrangement

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Abstract

Mass transport of the bulk of the analyte to the electrode and through the bioactive layer can be significantly improved by use of the nanoelectrode array and defined arrangement of protein film. This phenomenon has been studied by (i) atomic-force microscopy, (ii) electrochemical measurements of PSII activity, and (iii) digital simulations for an oriented monolayer of histidine-tagged photosystem II (PSII) immobilized on nitrilotriacetic acid (NTA)-modified gold electrodes. The output signal of the electrochemical biosensor is controlled by (i) mass transport from the bioactive layer to electrode and (ii) mass transport between the bulk of the analyte and the electrode. Mass transport through the bioactive layer was electrochemically studied for PSII self-assembled on gold screen-printed electrodes. A densely packed monolayer of PSII has a significant shielding effect toward the diffusion of redox mediator duroquinone (DQ). Mass transport to the planar electrode surface was improved by co-immobilization of bovine-serum albumin (BSA) as spacer biomolecule in the monolayer of PSII. Correlation between the electrochemical properties and surface arrangement of the resulting protein films was clearly observable and confirmed the improved mass-transport properties of structured enzyme monolayers. On the basis of this observation, the application of a bottom-up approach for improvement of electrode performance was proposed and digitally simulated for an infinite array of electrodes ranging in diameter from 50 nm to 5 μm. The nanoelectrode array, with the optimum time window selected for measurements, enables enhancement of mass transport between the bulk of the analyte and the macroelectrode by a factor of up to 50 in comparison with “classical” planar electrodes. Use of a time window enables minimization of crosstalk between individual electrodes in the array. The measurements require methods which suppress the double-layer capacity.

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Abbreviations

AuWE:

Gold working electrode

BSA:

Bovine serum albumin

Chl:

Chlorophyll

CMLH:

Nα,Nα-Bis(carboxymethyl)-l-lysine hydrate

DQ:

Tetramethyl-p-benzoquinone

DM:

Dodecylmaltoside

GA:

Glutaraldehyde

His-PSII:

Histidine-tagged photosystem II

LED:

Light-emitting diode

ME:

Macroelectrode consisting of an array of nanoelectrodes

MES:

2-(N-Morpholino)ethanesulfonic acid

NE:

Nanoelectrode

NTA:

Nitrilotriacetic acid

NTA-AuWE:

Gold working electrode modified with SAM layer of nitrilotriacetic acid

PEO:

1-Cyclohexyl-3-(2-morpholinoethyl)carbodiimidemetho-p-toluene sulfonate

PSII:

Photosystem II

PSII-NTA-AuWE:

His-PSII immobilized on NTA-AuWE

SAM:

Self-assembled monolayer

PB:

Phosphate buffer

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Acknowledgments

This work was supported by EU project Growth GRD1-2001-41831 “Microprotein”, by project 522/03/0659 of the Grant Agency of the Czech Republic, by project “IBIS” of the Czech Ministry of Industry and Trade, and by the COSMIC Project (ENEA Target Project on Biosensors and Bioelectronics).

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Authors and Affiliations

  1. Department of Biology, University of Jan Evangelista Purkyně, České mládeže 8, 400 96, Ústí nad Labem, Czech Republic

    J. Maly & Z. Stryhal

  2. Krejčí Engineering, Riegrova 318, 666 01, Tišnov, Czech Republic

    J. Krejci & R. Pilloton

  3. ENEA, SP061, Via Anguillarese 301, 00060 Santa Maria di Galeria, Roma, Italy

    M. Ilie

  4. Department of Microelectronics, CES Centre of Electrochemical Sensors, Joint Research Activity of BVT Technologies and University of Technology, FEEC, Údolní 53, 60200, Brno, Czech Republic

    L. Jakubka, K. Sameh & P. Steffan

  5. Institute of Microbiology, Academy of Sciences, Opatovický mlýn, 379 81, Třeboň, Czech Republic

    J. Masojídek

  6. Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, 595-8531, Osaka, Japan

    M. Sugiura

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  1. J. Maly
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Maly, J., Krejci, J., Ilie, M. et al. Monolayers of photosystem II on gold electrodes with enhanced sensor response—effect of porosity and protein layer arrangement. Anal Bioanal Chem 381, 1558–1567 (2005). https://doi.org/10.1007/s00216-005-3149-9

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  • DOI: https://doi.org/10.1007/s00216-005-3149-9

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