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Electrical contacting of glucose oxidase by DNA-templated polyaniline wires on surfaces
Electrochemistry Communications 6 (2004) 1057–1060 www.elsevier.com/locate/elecom
Electrical contacting of glucose oxidase by DNA-templated polyaniline wires on surfaces Lixin Shi, Yi Xiao, Itamar Willner
*
Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem 91904, Israel Received 21 July 2004; accepted 5 August 2004
Abstract Poly(aniline–aniline boronic acid) wires are generated on ds-DNA templates, and the resulting wires exhibit redox functions at neutral pH aqueous solutions. The association of flavin adenine dinucleotide (FAD) to the boronic acid ligand followed by the reconstitution of apo-glucose oxidase on the cofactor units yield an integrated enzyme-electrode where the biocatalyst reveals direct electrical contact with the electrode. 2004 Elsevier B.V. All rights reserved. Keywords: Poly(aniline–aniline boronic acid) wires; DNA template; Apo-glucose oxidase; Reconstitution; Biosensor
1. Introduction The electrical contacting of redox-enzymes with electrodes is a fundamental need for developing bioelectronic devices such as biosensor [1–5] or biofuel cell elements [6–8]. Recent advances in this area have reported on the effective electrical contacting of redoxenzymes with electrodes by the structural alignment of the biocatalysts on the electrodes by means of the reconstitution of the apo-enzyme or cofactor-modified conductive surfaces. Molecular relay units [9,10], conductive polymer matrices [11,12] or Au-nanoparticles [13] were used as electron transporting units that mediate electron transport between the enzyme redox-centers and the electrodes. Recently, single-wall carbon nanotubes were employed as nanowires that electrically contact glucose oxidase reconstituted on the ends of a cofactor-functionalized carbon nanotube [14]. Here, we wish to report on the generation of redox-active poly(aniline–aniline boronic acid) copolymer wires on *
Corresponding author. Tel.: +972 2 6585272; fax: +972 2 6527715. E-mail address: [email protected] (I. Willner).
1388-2481/$ - see front matter 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.elecom.2004.08.004
DNA templates. These wires are deposited on an electrode support by means of electrostatic interactions, and the polyaniline exhibits redox activity at neutral pH solutions. The reconstitution of glucose oxidase on the polyaniline wires leads to the electrical contacting of the enzyme with the electrode and its activation towards the bioelectrocatalyzed oxidation of glucose by means of the polyaniline nanowires. In fact, in a recent report, a polyaniline copolymer was employed for the electrochemical detection of dopamine [15]. Polyaniline is redox-active only in acidic aqueous solutions. It was demonstrated, however, that the generation of polyaniline blends that included negatively charged co-components such as sulfonic acids or polyacrylic acids lead to polyaniline that exhibited redox activity in neutral aqueous solutions [16–18]. Recently, it was demonstrated that H2O2 that was electrogenerated by an intercalator bound to double-stranded DNA linked to an electrode could oxidize aniline units associated with the DNA in the presence of horseradish peroxidase (HRP) [19]. The resulting polyaniline generated on the DNA template revealed redox functions in neutral pH solutions. The generation of polyaniline on
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the DNA templates was, however, demonstrated only on short double-stranded nucleic acid templates (25–30 bases) without any structural characterization of the polymer wires. Also, no functions of the polyaniline on the DNA template were demonstrated.
2. Experimental 2.1. Materials Apo-glucose oxidase (apo-GOx) was prepared by a modification of the reported method [20]. All other chemicals, including k-phage DNA (from Escherichia coli host strain GM 119), flavin adenine dinucleotide (FAD), horseradish peroxidase (HRP) (EC 1.11.1.7 Type II: from horseradish), glucose oxidase (GOx, EC 1.1.3.4 from Aspergillus niger), mercaptopropionic acid, aniline, 3-aminophenylboronic acid, and hydrogen peroxide were purchased from Sigma and used as supplied. Ultrapure water from Barnstead NANOpure Diamond system was used in all experiments. 2.2. Preparation of poly(aniline–aniline boronic acid) on DNA template The polymerization of (aniline–aniline boronic acid) was carried out in 0.01 M phosphate buffer (pH 4.3) that included aniline, 4.7 · 10 6 M, and 3-aminophenylboronic acid, 5.2 · 10 7 M, (9:1 molar ratio) in the presence of k-phage DNA (20 lg/ml). A 200 ll solution of HRP (1.5 mg/ml) was then added to this solution and continuously stirred. The reaction was initiated with the addition of H2O2 [21]. To avoid the inhibition of HRP due to excess amount of H2O2, a 0.3% solution of H2O2 (300 ll) was added dropwise (20 ll), and the mixture was stirred for 1 h at RT after which a dark green DNA–poly(aniline–aniline boronic acid) was formed. The formation of the polymer was confirmed by absorption spectroscopy and electrochemical means (vide infra). No polymer formation was detected upon the addition of H2O2/HRP to the aniline/3-amino phenylboronic acid at pH 4.3 in the absence of DNA. The resulting solution was dialyzed against the deionized water overnight to remove unreacted monomer.
rinsed again with water. The Au wires were soaked in mercaptopropionic acid solution, 0.1 M, for 2 h and then rinsed thoroughly with water. The resulting wires were incubated in the solution of poly(aniline–aniline boronic acid) solution (pH 7.0), for 2 h. The flavin adenine dinucleotide, FAD, cofactor units were covalently bound to the polymer-modified interface via phenylboronic acid ligand units. The polymer-functionalized Au wires were reacted with the FAD solution, 1 mM in 0.1 M phosphate buffer, pH 7.0, for 2 h. The polymer– FAD-functionalized Au wires were reacted with apoGox, 1 mg/ml in 0.1 M phosphate buffer, pH 7.0, for 5 h at room temperature, and then overnight at 4 C. A conventional three-electrode cell, consisting of the modified Au wire as working electrode, a glassy carbon auxiliary electrode isolated by a glass frit, and a saturated calomel reference electrode (SCE) connected to the working volume with a Luggin capillary, was used for the electrochemical measurements. All potentials are reported with respect to the SCE. Argon bubbling was used to remove oxygen from the solutions in the electrochemical cell. Cyclic voltammetry measurements were performed using an elelctorchemical analyzer (Eg&g model 283). The SEM measurement was performed on an FEI-Sirion HRSEM using voltages of 3 kV.
3. Results and discussions Fig. 1 shows the SEM image of the polymer-coated DNA. Long fibers (ca. 5–15 lm long) with a thickness of ca. 500 nm are observed. The length of the polymer wires is substantially lower than the length of the kphage DNA. This presumably originates from the partial acidic hydrolysis of the DNA template under the experimental polymerization conditions. The poly(ani-
2.3. Modification and measurements of electrodes Au wires (0.5 mm diameter) were used for modifications. The Au wires were purified by the treatment with a piranha solution (consisting of 70% concentrated sulfuric acid and 30% hydrogen peroxide, CAUTION: piranha solution reacts violently with most organic materials and must be handled with extreme care) for 20 min, and then thoroughly rinsed with pure water. The plate was then soaked in concentrated nitric acid for 5 min, and
Fig. 1. SEM image of the poly(aniline–aniline boronic acid) wires generated on the DNA template. The surface was coated with a Au/Pt layer (7–8 nm) to enhance the conductivity of the support.
L. Shi et al. / Electrochemistry Communications 6 (2004) 1057–1060
line–aniline boronic acid) wires were then reacted with flavin adenine dinucleotide (FAD) units that bind to the boronic acid ligands. The resulting cofactorfunctionalized polymer wires were electrostatically bound to a mercaptopropionic acid monolayer-functionalized Au-electrode, Scheme 1. The cyclic voltammogram of the polymer-modified electrode showed a quasi-reversible redox-wave at 0.43 V vs. SCE, corresponding to the FAD units, and a broad redox-wave at 0.26 V characteristic to polyaniline. The modified electrode represents a stable assembly, and the components are not washed off by rinsing with an aqueous buffer solution. Fig. 2(A) shows the cyclic voltammograms of the FAD redox-active unit associated with the polyaniline wires on the Auelectrode at variable scan rates. The anodic (or cathodic) peak currents show a linear dependence on the scan rate, consistent with a surface confined species. The Laviron analysis [22], Fig. 2(B), of the peak-to-peak separation at variable scan-rates yields an electron transfer rateconstant from the FAD units to the electrode that corresponds to 70 s 1. The interaction of the FAD-modified polymer-functionalized electrode with apo-glucose oxidase (apoGOx) resulted in the reconstitution of apo-protein on the FAD cofactor units. Several methods were applied to characterize the system. Coulometric analysis of the FAD redox-wave indicates a surface coverage of ca. 7 · 10 12 mole cm 2 of the cofactor units. Microgravi-
Scheme 1. Assembly of electrically contacted glucose oxidase (GOx) on a poly(aniline–aniline boronic acid) wire organized on a DNA template.
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Fig. 2. (A) Cyclic voltammograms of the FAD units linked to the boronate sites of the poly(aniline–aniline boronic acid) wires that are electrostatically assembled on the Au-electrode (surface area 0.25 cm2, roughness factor 1.3), at variable scan rates (V s 1): (a) 9, (b) 10, (c) 12, (d) 18, (e) 19 and (f) 20. (B) Laviron analysis showing the peak-to-peak separation of the FAD electrochemical response as a function of the logarithm of the scan rate. Experiments were performed in a phosphate buffer solution, pH 7.0, under Argon.
metric-quartz-crystal-microbalance measurements indicate a surface coverage of the polymer-templated DNA of ca. 4.2 · 10 8 gr cm 2, whereas the surface coverage of the enzyme is ca. 0.9 · 10 12 mole cm 2. Fig. 3(A) shows the cyclic voltammograms observed upon the bioelectrocatalyzed oxidation of different concentrations of glucose by the modified electrode. Clearly, electrocatalytic anodic currents as a result of the electrocatalyzed oxidation of glucose are observed. Control experiments reveal that the deposition of the poly(aniline–aniline boronic acid) wires that lack the FAD units on the monolayer-modified electrode, and the interaction of the electrode with native GOx do not lead to a bioelectrocatalytically active electrode. It should be noted that the native GOx glycoprotein binds to the boronic acid ligands of the polymer, but it lacks direct electrical communication with the electrode. In fact, the boronate-bound GOx glycoprotein exists in a biocatalytically active configuration, and upon the addition of ferrocene carboxylic acid, 1 · 10 3 M, to the modified electrode, the effective bioelectrocatalyzed oxidation of glucose occurs. Thus, the structural alignment of GOx on the polymer wires by the reconstitution process is essential to generate the electrically-contacted enzyme electrode. Fig. 3(B) shows the derived calibration curve that corresponds to the amperometric responses
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Preliminary studies indicate that similar redox-active poly(aniline–aniline boronic acid) wires for the electrical contacting of glucose oxidase may be generated with other negatively charged polyelectrolyte templates such as polystyrene sulfonate.
Acknowledgements This research is supported by the German-Israeli Foundation (GIF).
References [1] [2] [3] [4] [5] [6] [7] [8] [9] Fig. 3. (A) Cyclic voltammograms corresponding to the bioelectrocatalyzed oxidation of different concentrations of glucose by the integrated electrically contacted GOx-electrode (surface area 0.25 cm2, roughness factor 1.3): (a) 0 mM, (b) 5 mM, (c) 15 mM, (d) 25 mM and (e) 50 mM. (B) Calibration curve corresponds to the currents generated by the integrated GOx electrode at different glucose concentrations (at E = 0.5 V). Experiments were performed in a phosphate buffer solution, pH 7.0, under Argon.
of the functionalized electrode in the presence of different concentrations of glucose.
4. Conclusions In summary, the present study demonstrates that poly(aniline–aniline boronic acid) coatings on DNA templates generate redox-active polymer wires that electrically contact glucose oxidase reconstituted on the polymer wires with the electrode. The alignment of the protein on the polymer wires by means of the reconstitution process facilitates the electrical contacting process.
[10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20]
[21] [22]
I. Willner, Science 298 (2002) 2407. F.A. Armstrong, G.S. Wilson, Electrochim. Acta 41 (2000) 2623. A. Heller, Acc. Chem. Res. 23 (1990) 128. A. Heller, J. Phys. Chem. 96 (1992) 3579. I. Willner, E. Katz, Angew. Chem., Int. Ed. 39 (2000) 1180. E. Katz, I. Willner, A.B. Kotlyar, J. Electroanal. Chem. 479 (1999) 64. E. Katz, I. Willner, J. Am. Chem. Soc. 125 (2000) 6803. N. Mano, F. Mao, A. Heller, J. Am. Chem. Soc. 125 (2003) 6588. I. Willner, V. Heleg-Shabtai, R. Blonder, E. Katz, G.L. Tao, J. Am. Chem. Soc. 118 (1996) 10321. W. Schuhmann, T.J. Ohara, H.L. Schmidt, A. Heller, J. Am. Chem. Soc. 113 (1991) 1394. O.A. Raitman, E. Katz, A.F. Bu¨ckmann, I. Willner, J. Am. Chem. Soc. 124 (2002) 6487. O.A. Raitman, F. Patolsky, E. Katz, I. Willner, Chem. Commun. (2002) 1936. Y. Xiao, F. Patolsky, E. Katz, J.F. Hainfeld, I. Willner, Science 299 (2003) 1877. F. Patolsky, Y. Weizmann, I. Willner, Angew. Chem., Int. Ed. 43 (2004) 2113. B. Fabre, L. Tallebois, Chem. Commun. (2003) 2982. P.N. Bartlett, J.H. Wang, J. Chem. Soc., Faraday Trans. 92 (1996) 4137. P.N. Bartlett, E.N.K. Wallace, Phy. Chem. Chem. Phys. 3 (2001) 1491. P.N. Bartlett, E. Simon, Phys. Chem. Chem. Phys. 2 (2000) 2599. Y. Xiao, A.B. Kharitonov, F. Patolsky, Y. Weizmann, I. Willner, Chem. Commun. (2003) 1540. D.L. Morris, R.T. Buckler, in: J.J. Langone, H. Van Vunakis (Eds.), Methods in Enzymology, 92, Academic Press, Orlando, FL, 1983, pp. 413–417 (Part E). R. Nagarajan, W. Liu, J. Kumar, S.K. Tripathy, F.F. Baruno, L.A. Samuelson, Macromolecules 34 (2001) 3921. E. Laviron, J. Electroanal. Chem. 101 (1979) 19.
Electrical contacting of glucose oxidase by DNA-templated polyaniline wires on surfaces Lixin Shi, Yi Xiao, Itamar Willner
*
Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem 91904, Israel Received 21 July 2004; accepted 5 August 2004
Abstract Poly(aniline–aniline boronic acid) wires are generated on ds-DNA templates, and the resulting wires exhibit redox functions at neutral pH aqueous solutions. The association of flavin adenine dinucleotide (FAD) to the boronic acid ligand followed by the reconstitution of apo-glucose oxidase on the cofactor units yield an integrated enzyme-electrode where the biocatalyst reveals direct electrical contact with the electrode. 2004 Elsevier B.V. All rights reserved. Keywords: Poly(aniline–aniline boronic acid) wires; DNA template; Apo-glucose oxidase; Reconstitution; Biosensor
1. Introduction The electrical contacting of redox-enzymes with electrodes is a fundamental need for developing bioelectronic devices such as biosensor [1–5] or biofuel cell elements [6–8]. Recent advances in this area have reported on the effective electrical contacting of redoxenzymes with electrodes by the structural alignment of the biocatalysts on the electrodes by means of the reconstitution of the apo-enzyme or cofactor-modified conductive surfaces. Molecular relay units [9,10], conductive polymer matrices [11,12] or Au-nanoparticles [13] were used as electron transporting units that mediate electron transport between the enzyme redox-centers and the electrodes. Recently, single-wall carbon nanotubes were employed as nanowires that electrically contact glucose oxidase reconstituted on the ends of a cofactor-functionalized carbon nanotube [14]. Here, we wish to report on the generation of redox-active poly(aniline–aniline boronic acid) copolymer wires on *
Corresponding author. Tel.: +972 2 6585272; fax: +972 2 6527715. E-mail address: [email protected] (I. Willner).
1388-2481/$ - see front matter 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.elecom.2004.08.004
DNA templates. These wires are deposited on an electrode support by means of electrostatic interactions, and the polyaniline exhibits redox activity at neutral pH solutions. The reconstitution of glucose oxidase on the polyaniline wires leads to the electrical contacting of the enzyme with the electrode and its activation towards the bioelectrocatalyzed oxidation of glucose by means of the polyaniline nanowires. In fact, in a recent report, a polyaniline copolymer was employed for the electrochemical detection of dopamine [15]. Polyaniline is redox-active only in acidic aqueous solutions. It was demonstrated, however, that the generation of polyaniline blends that included negatively charged co-components such as sulfonic acids or polyacrylic acids lead to polyaniline that exhibited redox activity in neutral aqueous solutions [16–18]. Recently, it was demonstrated that H2O2 that was electrogenerated by an intercalator bound to double-stranded DNA linked to an electrode could oxidize aniline units associated with the DNA in the presence of horseradish peroxidase (HRP) [19]. The resulting polyaniline generated on the DNA template revealed redox functions in neutral pH solutions. The generation of polyaniline on
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L. Shi et al. / Electrochemistry Communications 6 (2004) 1057–1060
the DNA templates was, however, demonstrated only on short double-stranded nucleic acid templates (25–30 bases) without any structural characterization of the polymer wires. Also, no functions of the polyaniline on the DNA template were demonstrated.
2. Experimental 2.1. Materials Apo-glucose oxidase (apo-GOx) was prepared by a modification of the reported method [20]. All other chemicals, including k-phage DNA (from Escherichia coli host strain GM 119), flavin adenine dinucleotide (FAD), horseradish peroxidase (HRP) (EC 1.11.1.7 Type II: from horseradish), glucose oxidase (GOx, EC 1.1.3.4 from Aspergillus niger), mercaptopropionic acid, aniline, 3-aminophenylboronic acid, and hydrogen peroxide were purchased from Sigma and used as supplied. Ultrapure water from Barnstead NANOpure Diamond system was used in all experiments. 2.2. Preparation of poly(aniline–aniline boronic acid) on DNA template The polymerization of (aniline–aniline boronic acid) was carried out in 0.01 M phosphate buffer (pH 4.3) that included aniline, 4.7 · 10 6 M, and 3-aminophenylboronic acid, 5.2 · 10 7 M, (9:1 molar ratio) in the presence of k-phage DNA (20 lg/ml). A 200 ll solution of HRP (1.5 mg/ml) was then added to this solution and continuously stirred. The reaction was initiated with the addition of H2O2 [21]. To avoid the inhibition of HRP due to excess amount of H2O2, a 0.3% solution of H2O2 (300 ll) was added dropwise (20 ll), and the mixture was stirred for 1 h at RT after which a dark green DNA–poly(aniline–aniline boronic acid) was formed. The formation of the polymer was confirmed by absorption spectroscopy and electrochemical means (vide infra). No polymer formation was detected upon the addition of H2O2/HRP to the aniline/3-amino phenylboronic acid at pH 4.3 in the absence of DNA. The resulting solution was dialyzed against the deionized water overnight to remove unreacted monomer.
rinsed again with water. The Au wires were soaked in mercaptopropionic acid solution, 0.1 M, for 2 h and then rinsed thoroughly with water. The resulting wires were incubated in the solution of poly(aniline–aniline boronic acid) solution (pH 7.0), for 2 h. The flavin adenine dinucleotide, FAD, cofactor units were covalently bound to the polymer-modified interface via phenylboronic acid ligand units. The polymer-functionalized Au wires were reacted with the FAD solution, 1 mM in 0.1 M phosphate buffer, pH 7.0, for 2 h. The polymer– FAD-functionalized Au wires were reacted with apoGox, 1 mg/ml in 0.1 M phosphate buffer, pH 7.0, for 5 h at room temperature, and then overnight at 4 C. A conventional three-electrode cell, consisting of the modified Au wire as working electrode, a glassy carbon auxiliary electrode isolated by a glass frit, and a saturated calomel reference electrode (SCE) connected to the working volume with a Luggin capillary, was used for the electrochemical measurements. All potentials are reported with respect to the SCE. Argon bubbling was used to remove oxygen from the solutions in the electrochemical cell. Cyclic voltammetry measurements were performed using an elelctorchemical analyzer (Eg&g model 283). The SEM measurement was performed on an FEI-Sirion HRSEM using voltages of 3 kV.
3. Results and discussions Fig. 1 shows the SEM image of the polymer-coated DNA. Long fibers (ca. 5–15 lm long) with a thickness of ca. 500 nm are observed. The length of the polymer wires is substantially lower than the length of the kphage DNA. This presumably originates from the partial acidic hydrolysis of the DNA template under the experimental polymerization conditions. The poly(ani-
2.3. Modification and measurements of electrodes Au wires (0.5 mm diameter) were used for modifications. The Au wires were purified by the treatment with a piranha solution (consisting of 70% concentrated sulfuric acid and 30% hydrogen peroxide, CAUTION: piranha solution reacts violently with most organic materials and must be handled with extreme care) for 20 min, and then thoroughly rinsed with pure water. The plate was then soaked in concentrated nitric acid for 5 min, and
Fig. 1. SEM image of the poly(aniline–aniline boronic acid) wires generated on the DNA template. The surface was coated with a Au/Pt layer (7–8 nm) to enhance the conductivity of the support.
L. Shi et al. / Electrochemistry Communications 6 (2004) 1057–1060
line–aniline boronic acid) wires were then reacted with flavin adenine dinucleotide (FAD) units that bind to the boronic acid ligands. The resulting cofactorfunctionalized polymer wires were electrostatically bound to a mercaptopropionic acid monolayer-functionalized Au-electrode, Scheme 1. The cyclic voltammogram of the polymer-modified electrode showed a quasi-reversible redox-wave at 0.43 V vs. SCE, corresponding to the FAD units, and a broad redox-wave at 0.26 V characteristic to polyaniline. The modified electrode represents a stable assembly, and the components are not washed off by rinsing with an aqueous buffer solution. Fig. 2(A) shows the cyclic voltammograms of the FAD redox-active unit associated with the polyaniline wires on the Auelectrode at variable scan rates. The anodic (or cathodic) peak currents show a linear dependence on the scan rate, consistent with a surface confined species. The Laviron analysis [22], Fig. 2(B), of the peak-to-peak separation at variable scan-rates yields an electron transfer rateconstant from the FAD units to the electrode that corresponds to 70 s 1. The interaction of the FAD-modified polymer-functionalized electrode with apo-glucose oxidase (apoGOx) resulted in the reconstitution of apo-protein on the FAD cofactor units. Several methods were applied to characterize the system. Coulometric analysis of the FAD redox-wave indicates a surface coverage of ca. 7 · 10 12 mole cm 2 of the cofactor units. Microgravi-
Scheme 1. Assembly of electrically contacted glucose oxidase (GOx) on a poly(aniline–aniline boronic acid) wire organized on a DNA template.
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Fig. 2. (A) Cyclic voltammograms of the FAD units linked to the boronate sites of the poly(aniline–aniline boronic acid) wires that are electrostatically assembled on the Au-electrode (surface area 0.25 cm2, roughness factor 1.3), at variable scan rates (V s 1): (a) 9, (b) 10, (c) 12, (d) 18, (e) 19 and (f) 20. (B) Laviron analysis showing the peak-to-peak separation of the FAD electrochemical response as a function of the logarithm of the scan rate. Experiments were performed in a phosphate buffer solution, pH 7.0, under Argon.
metric-quartz-crystal-microbalance measurements indicate a surface coverage of the polymer-templated DNA of ca. 4.2 · 10 8 gr cm 2, whereas the surface coverage of the enzyme is ca. 0.9 · 10 12 mole cm 2. Fig. 3(A) shows the cyclic voltammograms observed upon the bioelectrocatalyzed oxidation of different concentrations of glucose by the modified electrode. Clearly, electrocatalytic anodic currents as a result of the electrocatalyzed oxidation of glucose are observed. Control experiments reveal that the deposition of the poly(aniline–aniline boronic acid) wires that lack the FAD units on the monolayer-modified electrode, and the interaction of the electrode with native GOx do not lead to a bioelectrocatalytically active electrode. It should be noted that the native GOx glycoprotein binds to the boronic acid ligands of the polymer, but it lacks direct electrical communication with the electrode. In fact, the boronate-bound GOx glycoprotein exists in a biocatalytically active configuration, and upon the addition of ferrocene carboxylic acid, 1 · 10 3 M, to the modified electrode, the effective bioelectrocatalyzed oxidation of glucose occurs. Thus, the structural alignment of GOx on the polymer wires by the reconstitution process is essential to generate the electrically-contacted enzyme electrode. Fig. 3(B) shows the derived calibration curve that corresponds to the amperometric responses
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L. Shi et al. / Electrochemistry Communications 6 (2004) 1057–1060
Preliminary studies indicate that similar redox-active poly(aniline–aniline boronic acid) wires for the electrical contacting of glucose oxidase may be generated with other negatively charged polyelectrolyte templates such as polystyrene sulfonate.
Acknowledgements This research is supported by the German-Israeli Foundation (GIF).
References [1] [2] [3] [4] [5] [6] [7] [8] [9] Fig. 3. (A) Cyclic voltammograms corresponding to the bioelectrocatalyzed oxidation of different concentrations of glucose by the integrated electrically contacted GOx-electrode (surface area 0.25 cm2, roughness factor 1.3): (a) 0 mM, (b) 5 mM, (c) 15 mM, (d) 25 mM and (e) 50 mM. (B) Calibration curve corresponds to the currents generated by the integrated GOx electrode at different glucose concentrations (at E = 0.5 V). Experiments were performed in a phosphate buffer solution, pH 7.0, under Argon.
of the functionalized electrode in the presence of different concentrations of glucose.
4. Conclusions In summary, the present study demonstrates that poly(aniline–aniline boronic acid) coatings on DNA templates generate redox-active polymer wires that electrically contact glucose oxidase reconstituted on the polymer wires with the electrode. The alignment of the protein on the polymer wires by means of the reconstitution process facilitates the electrical contacting process.
[10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20]
[21] [22]
I. Willner, Science 298 (2002) 2407. F.A. Armstrong, G.S. Wilson, Electrochim. Acta 41 (2000) 2623. A. Heller, Acc. Chem. Res. 23 (1990) 128. A. Heller, J. Phys. Chem. 96 (1992) 3579. I. Willner, E. Katz, Angew. Chem., Int. Ed. 39 (2000) 1180. E. Katz, I. Willner, A.B. Kotlyar, J. Electroanal. Chem. 479 (1999) 64. E. Katz, I. Willner, J. Am. Chem. Soc. 125 (2000) 6803. N. Mano, F. Mao, A. Heller, J. Am. Chem. Soc. 125 (2003) 6588. I. Willner, V. Heleg-Shabtai, R. Blonder, E. Katz, G.L. Tao, J. Am. Chem. Soc. 118 (1996) 10321. W. Schuhmann, T.J. Ohara, H.L. Schmidt, A. Heller, J. Am. Chem. Soc. 113 (1991) 1394. O.A. Raitman, E. Katz, A.F. Bu¨ckmann, I. Willner, J. Am. Chem. Soc. 124 (2002) 6487. O.A. Raitman, F. Patolsky, E. Katz, I. Willner, Chem. Commun. (2002) 1936. Y. Xiao, F. Patolsky, E. Katz, J.F. Hainfeld, I. Willner, Science 299 (2003) 1877. F. Patolsky, Y. Weizmann, I. Willner, Angew. Chem., Int. Ed. 43 (2004) 2113. B. Fabre, L. Tallebois, Chem. Commun. (2003) 2982. P.N. Bartlett, J.H. Wang, J. Chem. Soc., Faraday Trans. 92 (1996) 4137. P.N. Bartlett, E.N.K. Wallace, Phy. Chem. Chem. Phys. 3 (2001) 1491. P.N. Bartlett, E. Simon, Phys. Chem. Chem. Phys. 2 (2000) 2599. Y. Xiao, A.B. Kharitonov, F. Patolsky, Y. Weizmann, I. Willner, Chem. Commun. (2003) 1540. D.L. Morris, R.T. Buckler, in: J.J. Langone, H. Van Vunakis (Eds.), Methods in Enzymology, 92, Academic Press, Orlando, FL, 1983, pp. 413–417 (Part E). R. Nagarajan, W. Liu, J. Kumar, S.K. Tripathy, F.F. Baruno, L.A. Samuelson, Macromolecules 34 (2001) 3921. E. Laviron, J. Electroanal. Chem. 101 (1979) 19.