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Author: Wollenberger, U. ×

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Now showing 1 - 10 of 41
  • Publication
    The electrically wired molybdenum domain of human sulfite oxidase is bioelectrocatalytically active
    ( 2015)
    Spricigo, R.
    ;
    Leimkühler, S.
    ;
    Gorton, L.
    ;
    Scheller, F.W.
    ;
    Wollenberger, U.
    We report electron transfer between the catalytic molybdenum cofactor (Moco) domain of human sulfite oxidase (hSO) and electrodes through a poly(vinylpyridine)-bound [osmium(N,N'-methyl-2,2'-biimidazole)3]2+/3+ complex as the electron-transfer mediator. The biocatalyst was immobilized in this low-potential redox polymer on a carbon electrode. Upon the addition of sulfite to the immobilized separate Moco domain, the generation of a significant catalytic current demonstrated that the catalytic center is effectively wired and active. The bioelectrocatalytic current of the wired separate catalytic domain reached 25 % of the signal of the wired full molybdoheme enzyme hSO, in which the heme b5 is involved in the electron-transfer pathway. This is the first report on a catalytically active wired molybdenum cofactor domain. The formal potential of this electrochemical mediator is between the potentials of the two cofactors of hSO, and as hSO can occupy several conformations in the polymer matrix, it is imaginable that electron transfer from the catalytic site to the electrode through the osmium center occurs for the hSO molecules in which the Moco domain is sufficiently accessible. The observation of catalytic oxidation currents at low potentials is favorable for applications in bioelectronic devices.
  • Publication
    Surface-tuned electron transfer and electrocatalysis of hexameric tyrosine-coordinated heme protein
    ( 2015)
    Peng, L.
    ;
    Utesch, T.
    ;
    Yarman, A.
    ;
    Jeoung, J.-H.
    ;
    Steinborn, S.
    ;
    Dobbek, H.
    ;
    Mroginski, M.A.
    ;
    Tanne, J.
    ;
    Wollenberger, U.
    ;
    Scheller, F.W.
    Molecular modeling, electrochemical methods, and quartz crystal microbalance were used to characterize immobilized hexameric tyrosine-coordinated heme protein (HTHP) on bare carbon or on gold electrodes modified with positively and negatively charged self-assembled monolayers (SAMs), respectively. HTHP binds to the positively charged surface but no direct electron transfer (DET) is found due to the long distance of the active sites from the electrode surfaces. At carboxyl-terminated surfaces, the neutrally charged bottom of HTHP can bind to the SAM. For this "disc" orientation all six hemes are close to the electrode and their direct electron transfer should be efficient. HTHP on all negatively charged SAMs showed a quasi-reversible redox behavior with rate constant ks values between 0.93 and 2.86 s-1 and apparent formal potentials E(Formula presented.) between -131.1 and -249.1 mV. On the MUA/MU-modified electrode, the maximum surface concentration corresponds to a complete monolayer of the hexameric HTHP in the disc orientation. HTHP electrostatically immobilized on negatively charged SAMs shows electrocatalysis of peroxide reduction and enzymatic oxidation of NADH. Heme theme: Electrochemistry and a quartz crystal microbalance were used to characterize hexameric tyrosine-coordinated heme protein (HTHP) immobilized on bare carbon or on Au electrodes modified by differently charged self-assembled monolayers. Experimental results are in line with theoretical calculations; HTHP at negatively charged surfaces shows both direct electron transfer of the heme centers and bioelectrocatalysis.
  • Publication
    Characterization of the enhanced peroxidatic activity of amyloid v peptide-hemin complexes towards neurotransmitters
    ( 2014)
    Neumann, B.
    ;
    Yarman, A.
    ;
    Wollenberger, U.
    ;
    Scheller, F.
    Binding of heme to the amyloid peptides A40/42 is thought to be an initial step in the development of symptoms in the early stages of Alzheimer's disease by enhancing the intrinsic peroxidatic activity of heme. We found considerably higher acceleration of the reaction for the physiologically relevant neurotransmitters dopamine and serotonin than reported earlier for the artificial substrate 3,3,5,5-tetramethylbenzidine (TMB). Thus, the binding of hemin to A peptides might play an even more crucial role in the early stages of Alzheimer's disease than deduced from these earlier results. To mimic complex formation, a new surface architecture has been developed: The interaction between the truncated amyloid peptide A1-16 and hemin immobilized on an aminohexanethiol spacer on a gold electrode has been analyzed by cyclic voltammetry. The resulting complex has a redox pair with a 25 mV more cathodic formal potential than hemin alone.
  • Publication
    Cytochrome P450: Electron transfer and sensors
    ( 2014)
    Scheller, F.W.
    ;
    Yarmana, A.
    ;
    Wollenberger, U.
    Electrochemical methods allow to characterize the reaction mechanism of the cytochrome P450 (CYP) enzymes by observing the electron transfer in real time. According to the number of publications on protein electrochemistry, CYP enzymes have the third position after glucose oxidase and cytochrome c. Sensors for drug monitoring using different CYP enzymes are appropriate tools since CYP enzymes act on more than 90 percent of all drugs currently on the market. For sensor development the efficiency of coupling the biocatalytic systems with the electrode is the crucial step. Furthermore for screening drugs, and for assessment of toxicity and prediction of drug clearance, the distribution of CYP isoenzymes and polymorphic enzymes would be of high clinical relevance.
  • Publication
    Surface modification with thermoresponsive polymer brushes for a switchable electrochemical sensor
    ( 2014)
    Comminges, C.
    ;
    Frasca, S.
    ;
    Sütterlin, M.
    ;
    Wischerhoff, E.
    ;
    Laschewsky, A.
    ;
    Wollenberger, U.
    Elaboration of switchable surfaces represents an interesting way for the development of a new generation of electrochemical sensors. In this paper, a method for growing thermoresponsive polymer brushes from a gold surface pre-modified with polyethyleneimine (PEI), subsequent layer-by-layer polyelectrolyte assembly and adsorption of a charged macroinitiator is described. We propose an easy method for monitoring the coil-to-globule phase transition of the polymer brush using an electrochemical quartz crystal microbalance with dissipation (E-QCM-D). The surface of these polymer modified electrodes shows reversible switching from the swollen to the collapsed state with temperature. As demonstrated from E-QCM-D measurements using an original signal processing method, the switch is operating in three reversible steps related to different interfacial viscosities. Moreover, it is shown that the one electron oxidation of ferrocene carboxylic acid is dramatically affected by the change from the swollen to the collapsed state of the polymer brush, showing a spectacular 86% decrease of the charge transfer resistance between the two states.
  • Publication
    Sensors based on cytochrome P450 and CYP mimicking systems
    ( 2013)
    Yarman, A.
    ;
    Wollenberger, U.
    ;
    Scheller, F.W.
    Cytochrome P450 enzymes (CYPs) act on more than 90 percent of all drugs currently on the market. The catalytic cycle requires electron supply to the heme iron in the presence of oxygen. Electrochemistry allows to characterise the reaction mechanism of these redox enzymes by observing the electron transfer in real time. According to the number of publications on protein electrochemistry CYP has the third position after glucose oxidase and cytochrome c. CYP based enzyme electrodes for the quantification of drugs, metabolites or pesticides have been developed using different iso-enzymes. A crucial step in the sensor development is the efficiency of coupling the biocatalytic systems with the electrode is. In the 1970s the direct electron transfer of heme and heme peptides called microperoxidases (MPs) was used as model of oxidoreductases. They exhibit a broad substrate spectrum including hydroxylation of selected aromatic substrates, demethylation and epoxidation by means o f hydrogen peroxide. It overlaps with that of P450 making heme and MPs to alternate recognition elements in biosensors for the detection of typical CYP substrates. In these enzyme electrodes the signal is generated by the conversion of all substrates thus representing in complex media an overall parameter. By combining the biocatalytic substrate conversion with selective binding to a molecularly imprinted polymer layer the specificity has been improved. Here we discuss different approaches of biosensors based on CYP, microperoxidases and catalytically active MIPs and discuss their potential as recognition elements in biosensors. The performance of these sensors and their further development are discussed.
  • Publication
    The aromatic peroxygenase from Marasmius rutola - a new enzyme for biosensor applications
    ( 2012)
    Yarman, A.
    ;
    Gröbe, G.
    ;
    Neumann, B.
    ;
    Kinne, M.
    ;
    Gajovic-Eichelmann, N.
    ;
    Wollenberger, U.
    ;
    Hofrichter, M.
    ;
    Ullrich, R.
    ;
    Scheibner, K.
    ;
    Scheller, F.W.
    The aromatic peroxygenase (APO; EC 1.11.2.1) from the agraric basidomycete Marasmius rotula (MroAPO) immobilized at the chitosan-capped gold-nanoparticle-modified glassy carbon electrode displayed a pair of redox peaks with a midpoint potential of -278.5 mV vs. AgCl/AgCl (1 M KCl) for the Fe2+/Fe3+ redox couple of the heme-thiolate-containing protein. MroAPO oxidizes aromatic substrates such as aniline, p-aminophenol, hydroquinone, resorcinol, catechol, and paracetamol by means of hydrogen peroxide. The substrate spectrum overlaps with those of cytochrome P450s and plant peroxidases which are relevant in environmental analysis and drug monitoring. In M. rotula peroxygenase-based enzyme electrodes, the signal is generated by the reduction of electrode-active reaction products (e.g., p-benzoquinone and p-quinoneimine) with electro-enzymatic recycling of the analyte. In these enzyme electrodes, the signal reflects the conversion of all substrates thus representing an overa ll parameter in complex media. The performance of these sensors and their further development are discussed.
  • Publication
    Enzyme electrode for aromatic compounds exploiting the catalytic activities of microperoxidase-11
    ( 2011)
    Yarman, A.
    ;
    Badalyan, A.
    ;
    Gajovic-Eichelmann, N.
    ;
    Wollenberger, U.
    ;
    Scheller, F.W.
    Microperoxidase-11 (MP-11) which has been immobilised in a matrix of chitosan-embedded gold nanoparticles on the surface of a glassy carbon electrode catalyzes the conversion of aromatic substances. This peroxide-dependent catalysis of microperoxidase has been applied in an enzyme electrode for the first time to indicate aromatic compounds such as aniline, 4-fluoroaniline, catechol and p-aminophenol. The electrode signal is generated by the cathodic reduction of the quinone or quinoneimine which is formed in the presence of both MP-11 and peroxide from the substrate. The same sensor principle will be extended to aromatic drugs.
  • Publication
    Bioelectrocatalysis by Microperoxidase-11 in a multilayer architecture of chitosan embedded gold nanoparticles
    ( 2011)
    Yarman, A.
    ;
    Nagel, T.
    ;
    Gajovic-Eichelmann, N.
    ;
    Fischer, A.
    ;
    Wollenberger, U.
    ;
    Scheller, F.W.
    We report on the redox behaviour of the microperoxidase-11 (MP-11) which has been electrostatically immobilized in a matrix of chitosan-embedded gold nanoparticles on the surface of a glassy carbon electrode. MP-11 contains a covalently bound heme c as the redox active group that exchanges electrons with the electrode via the gold nanoparticles. Electroactive surface concentration of MP-11 at high scan rate is between 350±50pmol cm-2, which reflects a multilayer process. The formal potential (E°') of MP-11 in the gold nanoparticles-chitosan film was estimated to be -(267.7±2.9) mV at pH7.0. The heterogeneous electron transfer rate constant (ks) starts at 1.21s-1 and levels off at 6.45s-1 in the scan rate range from 0.1 to 2.0V s-1. Oxidation and reduction of MP-11 by hydrogen peroxide and superoxide, respectively have been coupled to the direct electron transfer of MP-11.
  • Publication
    Direct electron transfer of Agrocybe aegerita peroxygenase at electrodes modified with chitosan-capped Au nanoparticles and its bioelectrocatalysis to aniline
    ( 2011)
    Wu, Y.
    ;
    Wollenberger, U.
    ;
    Hofrichter, M.
    ;
    Ullrich, R.
    ;
    Scheibner, K.
    ;
    Scheller, F.W.
    Three different sizes of chitosan-capped Au nanoparticles were synthesized and were used to incorporate Agrocybe aegerita peroxygenase (AaeAPO) onto the surface of glassy carbon electrode. The direct electron transfer of AaeAPO was achieved in all films. The highest amount of electroactive enzyme and highest electron transfer rate constant ks of AaeAPO were obtained in the film with the smallest size of chitosan-capped Au nanoparticles. In anaerobic solutions, quasi-reversible oxidation and reduction are obtained with a formal potential of -0.280 V vs. Ag/AgCl 1 M KCl in 100 mM (pH 7.0) PBS at scan rate of 1 V s-1. Bioelectrocatalytic reduction currents can be obtained with the AaeAPO-modified electrode on addition of hydrogen peroxide. This reaction was suppressed when sodium azide, an inhibitor of AaeAPO, was present. Furthermore, the peroxide-dependent conversion of aniline was characterized and it was found that a polymer product via p-aminophenol is formed. And the AaeAPO biosensor was applied to determine aniline and p-aminophenol.