TY - JOUR A1 - Muschallik, Lukas A1 - Molinnus, Denise A1 - Bongaerts, Johannes A1 - Pohl, Martina A1 - Wagner, Torsten A1 - Schöning, Michael Josef A1 - Siegert, Petra A1 - Selmer, Thorsten T1 - (R,R)-Butane-2,3-diol Dehydrogenase from Bacillus clausii DSM 8716T: Cloning and Expression of the bdhA-Gene, and Initial Characterization of Enzyme JF - Journal of Biotechnology N2 - The gene encoding a putative (R,R)-butane-2,3-diol dehydrogenase (bdhA) from Bacillus clausii DSM 8716T was isolated, sequenced and expressed in Escherichia coli. The amino acid sequence of the encoded protein is only distantly related to previously studied enzymes (identity 33–43%) and exhibited some uncharted peculiarities. An N-terminally StrepII-tagged enzyme variant was purified and initially characterized. The isolated enzyme catalyzed the (R)-specific oxidation of (R,R)- and meso-butane-2,3-diol to (R)- and (S)-acetoin with specific activities of 12 U/mg and 23 U/mg, respectively. Likewise, racemic acetoin was reduced with a specific activity of up to 115 U/mg yielding a mixture of (R,R)- and meso-butane-2,3-diol, while the enzyme reduced butane-2,3-dione (Vmax 74 U/mg) solely to (R,R)-butane-2,3-diol via (R)-acetoin. For these reactions only activity with the co-substrates NADH/NAD+ was observed. The enzyme accepted a selection of vicinal diketones, α-hydroxy ketones and vicinal diols as alternative substrates. Although the physiological function of the enzyme in B. clausii remains elusive, the data presented herein clearly demonstrates that the encoded enzyme is a genuine (R,R)-butane-2,3-diol dehydrogenase with potential for applications in biocatalysis and sensor development. Y1 - 2017 U6 - http://dx.doi.org/10.1016/j.jbiotec.2017.07.020 SN - 0168-1656 VL - 258 SP - 41 EP - 50 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Miyamoto, Ko-ichiro A1 - Hayashi, Kosuke A1 - Sakamoto, Azuma A1 - Werner, Frederik A1 - Wagner, Torsten A1 - Schöning, Michael Josef A1 - Yoshinobu, Tatsuo T1 - A high-Q resonance-mode measurement of EIS capacitive sensor by elimination of series resistance JF - Sensor and Actuators B: Chemical N2 - An EIS capacitive sensor is a semiconductor-based potentiometric sensor, which is sensitive to the ion concentration or pH value of the solution in contact with the sensing surface. To detect a small change in the ion concentration or pH, a small capacitance change must be detected. Recently, a resonance-mode measurement was proposed, in which an inductor was connected to the EIS capacitive sensor and the resonant frequency was correlated with the pH value. In this study, the Q factor of the resonant circuit was enhanced by canceling the internal resistance of the reference electrode and the internal resistance of the inductor coil with the help of a bypass capacitor and a negative impedance converter, respectively. 1% variation of the signal in the developed system corresponded to a pH change of 3.93 mpH, which was about 1/12 of the conventional method, suggesting a better performance in detection of a small pH change. KW - Negative impedance convertor KW - Resonance-mode measurement KW - Chemical sensor KW - EIS capacitive sensor Y1 - 2017 U6 - http://dx.doi.org/10.1016/j.snb.2017.03.002 SN - 0925-4005 VL - 248 SP - 1006 EP - 1010 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Honarvarfard, Elham A1 - Gamella, Maria A1 - Poghossian, Arshak A1 - Schöning, Michael Josef A1 - Katz, Evgeny T1 - An enzyme-based reversible Controlled NOT (CNOT) logic gate operating on a semiconductor transducer JF - Applied Materials Today N2 - An enzyme-based biocatalytic system mimicking operation of a logically reversible Controlled NOT (CNOT) gate has been interfaced with semiconductor electronic transducers. Electrolyte–insulator–semiconductor (EIS) structures have been used to transduce chemical changes produced by the enzyme system to an electronically readable capacitive output signal using field-effect features of the EIS device. Two enzymes, urease and esterase, were immobilized on the insulating interface of EIS structure producing local pH changes performing XOR logic operation controlled by various combinations of the input signals represented by urea and ethyl butyrate. Another EIS transducer was functionalized with esterase only, thus performing Identity (ID) logic operation for the ethyl butyrate input. Both semiconductor devices assembled in parallel operated as a logically reversible CNOT gate. The present system, despite its simplicity, demonstrated for the first time logically reversible function of the enzyme system transduced electronically with the semiconductor devices. The biomolecular realization of a CNOT gate interfaced with semiconductors is promising for integration into complex biomolecular networks and future biosensor/biomedical applications. KW - Electrolyte–insulator–semiconductor KW - Capacitive field-effect KW - CNOT KW - XOR KW - Enzyme logic gate Y1 - 2017 U6 - http://dx.doi.org/10.1016/j.apmt.2017.08.003 SN - 2352-9407 VL - 9 SP - 266 EP - 270 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Scholl, Fabio A1 - Morais, Paulo A1 - Gabriel, Rayla A1 - Schöning, Michael Josef A1 - Siqueira, Jose Roberto, Jr. A1 - Caseli, Luciano T1 - Carbon nanotubes arranged as smart interfaces in lipid Langmuir-Blodgett films enhancing the enzymatic properties of penicillinase for biosensing applications JF - Applied Materials & Interfaces N2 - In this paper, carbon nanotubes (CNTs) were incorporated in penicillinase-phospholipid Langmuir and Langmuir–Blodgett (LB) films to enhance the enzyme catalytic properties. Adsorption of the penicillinase and CNTs at dimyristoylphosphatidic acid (DMPA) monolayers at the air–water interface was investigated by surface pressure–area isotherms, vibrational spectroscopy, and Brewster angle microscopy. The floating monolayers were transferred to solid supports through the LB technique, forming mixed DMPA-CNTs-PEN films, which were investigated by quartz crystal microbalance, vibrational spectroscopy, and atomic force microscopy. Enzyme activity was studied with UV–vis spectroscopy and the feasibility of the supramolecular device nanostructured as ultrathin films were essayed in a capacitive electrolyte–insulator–semiconductor (EIS) sensor device. The presence of CNTs in the enzyme–lipid LB film not only tuned the catalytic activity of penicillinase but also helped conserve its enzyme activity after weeks, showing increased values of activity. Viability as penicillin sensor was demonstrated with capacitance/voltage and constant capacitance measurements, exhibiting regular and distinctive output signals over all concentrations used in this work. These results may be related not only to the nanostructured system provided by the film, but also to the synergism between the compounds on the active layer, leading to a surface morphology that allowed a fast analyte diffusion because of an adequate molecular accommodation, which also preserved the penicillinase activity. This work therefore demonstrates the feasibility of employing LB films composed of lipids, CNTs, and enzymes as EIS devices for biosensing applications. Y1 - 2017 U6 - http://dx.doi.org/10.1021/acsami.7b08095 SN - 1944-8252 VL - 9 IS - 36 SP - 31054 EP - 31066 PB - ACS CY - Washington ER - TY - JOUR A1 - Molinnus, Denise A1 - Poghossian, Arshak A1 - Keusgen, Michael A1 - Katz, Evgeny A1 - Schöning, Michael Josef T1 - Coupling of Biomolecular Logic Gates with Electronic Transducers: From Single Enzyme Logic Gates to Sense/Act/Treat Chips JF - Electroanalysis N2 - The integration of biomolecular logic principles with electronic transducers allows designing novel digital biosensors with direct electrical output, logically triggered drug-release, and closed-loop sense/act/treat systems. This opens new opportunities for advanced personalized medicine in the context of theranostics. In the present work, we will discuss selected examples of recent developments in the field of interfacing enzyme logic gates with electrodes and semiconductor field-effect devices. Special attention is given to an enzyme OR/Reset logic gate based on a capacitive field-effect electrolyte-insulator-semiconductor sensor modified with a multi-enzyme membrane. Further examples are a digital adrenaline biosensor based on an AND logic gate with binary YES/NO output and an integrated closed-loop sense/act/treat system comprising an amperometric glucose sensor, a hydrogel actuator, and an insulin (drug) sensor. Y1 - 2017 U6 - http://dx.doi.org/10.1002/elan.201700208 SN - 1521-4109 VL - 29 IS - 8 SP - 1840 EP - 1849 PB - Wiley CY - Weinheim ER - TY - CHAP A1 - Molinnus, Denise A1 - Hardt, Gabriel A1 - Käver, Larissa A1 - Willenberg, Holger S. A1 - Poghossian, Arshak A1 - Keusgen, Michael A1 - Schöning, Michael Josef T1 - Detection of Adrenaline Based on Bioelectrocatalytical System to Support Tumor Diagnostic Technology T2 - MDPI Proceedings Y1 - 2017 U6 - http://dx.doi.org/10.3390/proceedings1040506 ER - TY - JOUR A1 - Sousa, Marcos A. M. A1 - Siqueira, Jose R. Jr. A1 - Vercik, Andres A1 - Schöning, Michael Josef A1 - Oliveira, Osvaldo N. Jr. T1 - Determining the optimized layer-by-layer film architecture with dendrimer/carbon nanotubes for field-effect sensors JF - IEEE Sensors Journal N2 - The capacitive electrolyte–insulator–semiconductor (EIS) structure is a typical device based on a field-effect sensor platform. With a simple silicon-based structure, EIS have been useful for several sensing applications, especially with incorporation of nanostructured films to modulate the ionic transport and the flat-band potential. In this paper, we report on ion transport and changes in flat-band potential in EIS sensors made with layer-by-layer films containing poly(amidoamine) (PAMAM) dendrimer and single-walled carbon nanotubes (SWNTs) adsorbed on p-Si/SiO 2 /Ta 2 O 5 chips with an Al ohmic contact. The impedance spectra were fitted using an equivalent circuit model, from which we could determine parameters such as the double-layer capacitance. This capacitance decreased with the number of bilayers owing to space charge accumulated at the electrolyte–insulator interface, up to three PAMAM/SWNTs bilayers, after which it stabilized. The charge-transfer resistance was also minimum for three bilayers, thus indicating that this is the ideal architecture for an optimized EIS performance. The understanding of the influence of nanostructures and the fine control of operation parameters pave the way for optimizing the design and performance of new EIS sensors. Y1 - 2017 U6 - http://dx.doi.org/10.1109/JSEN.2017.2653238 SN - 1558-1748 VL - 17 IS - 6 SP - 1735 EP - 1740 PB - IEEE CY - New York ER - TY - JOUR A1 - Röhlen, Desiree A1 - Pilas, Johanna A1 - Schöning, Michael Josef A1 - Selmer, Thorsten T1 - Development of an amperometric biosensor platform for the combined determination of l-Malic, Fumaric, and l-Aspartic acid JF - Applied Biochemistry and Biotechnology N2 - Three amperometric biosensors have been developed for the detection of L-malic acid, fumaric acid, and L -aspartic acid, all based on the combination of a malate-specific dehydrogenase (MDH, EC 1.1.1.37) and diaphorase (DIA, EC 1.8.1.4). The stepwise expansion of the malate platform with the enzymes fumarate hydratase (FH, EC 4.2.1.2) and aspartate ammonia-lyase (ASPA, EC 4.3.1.1) resulted in multi-enzyme reaction cascades and, thus, augmentation of the substrate spectrum of the sensors. Electrochemical measurements were carried out in presence of the cofactor β-nicotinamide adenine dinucleotide (NAD+) and the redox mediator hexacyanoferrate (III) (HCFIII). The amperometric detection is mediated by oxidation of hexacyanoferrate (II) (HCFII) at an applied potential of + 0.3 V vs. Ag/AgCl. For each biosensor, optimum working conditions were defined by adjustment of cofactor concentrations, buffer pH, and immobilization procedure. Under these improved conditions, amperometric responses were linear up to 3.0 mM for L-malate and fumarate, respectively, with a corresponding sensitivity of 0.7 μA mM−1 (L-malate biosensor) and 0.4 μA mM−1 (fumarate biosensor). The L-aspartate detection system displayed a linear range of 1.0–10.0 mM with a sensitivity of 0.09 μA mM−1. The sensor characteristics suggest that the developed platform provides a promising method for the detection and differentiation of the three substrates. Y1 - 2017 U6 - http://dx.doi.org/10.1007/s12010-017-2578-1 SN - 1559-0291 VL - 183 SP - 566 EP - 581 PB - Springer CY - Berlin ER - TY - JOUR A1 - Dantism, Shahriar A1 - Takenaga, Shoko A1 - Wagner, Torsten A1 - Wagner, Patrick A1 - Schöning, Michael Josef T1 - Differential imaging of the metabolism of bacteria and eukaryotic cells based on light-addressable potentiometric sensors JF - Electrochimica Acta N2 - A light-addressable potentiometric sensor (LAPS) is a field-effect-based potentiometric sensor with an electrolyte/insulator/semiconductor (EIS) structure, which is able to monitor analyte concentrations of (bio-)chemical species in aqueous solutions in a spatially resolved way. Therefore, it is also an appropriate tool to record 2D-chemical images of concentration variations on the sensor surface. In the present work, two differential, LAPS-based measurement principles are introduced to determine the metabolic activity of Escherichia coli (E. coli) K12 and Chinese hamster ovary (CHO) cells as test microorganisms. Hereby, we focus on i) the determination of the extracellular acidification rate (ΔpH/min) after adding glucose solutions to the cell suspensions; and ii) recording the amplitude increase of the photocurrent (Iph) related to the produced acids from E. coli K12 bacteria and CHO cells on the sensor surface by 2D-chemical imaging. For this purpose, 3D-printed multi-chamber structures were developed and mounted on the planar sensor-chip surface to define four independent compartments, enabling differential measurements with varying cell concentrations. The differential concept allows eliminating unwanted drift effects and, with the four-chamber structures, measurements on the different cell concentrations were performed simultaneously, thus reducing also the overall measuring time. Y1 - 2017 U6 - http://dx.doi.org/10.1016/j.electacta.2017.05.196 SN - 0013-4686 VL - 246 SP - 234 EP - 241 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Gamella, Maria A1 - Zakharchenko, Andrey A1 - Guz, Nataliia A1 - Masi, Madeline A1 - Minko, Sergiy A1 - Kolpashchikov, Dmitry M. A1 - Iken, Heiko A1 - Poghossian, Arshak A1 - Schöning, Michael Josef A1 - Katz, Evgeny T1 - DNA computing system activated by electrochemically triggered DNA realease from a polymer-brush-modified electrode array JF - Electroanalysis N2 - An array of four independently wired indium tin oxide (ITO) electrodes was used for electrochemically stimulated DNA release and activation of DNA-based Identity, AND and XOR logic gates. Single-stranded DNA molecules were loaded on the mixed poly(N,N-dimethylaminoethyl methacrylate) (PDMAEMA)/poly(methacrylic acid) (PMAA) brush covalently attached to the ITO electrodes. The DNA deposition was performed at pH 5.0 when the polymer brush is positively charged due to protonation of tertiary amino groups in PDMAEMA, thus resulting in electrostatic attraction of the negatively charged DNA. By applying electrolysis at −1.0 V(vs. Ag/AgCl reference) electrochemical oxygen reduction resulted in the consumption of hydrogen ions and local pH increase near the electrode surface. The process resulted in recharging the polymer brush to the negative state due to dissociation of carboxylic groups of PMAA, thus repulsing the negatively charged DNA and releasing it from the electrode surface. The DNA release was performed in various combinations from different electrodes in the array assembly. The released DNA operated as input signals for activation of the Boolean logic gates. The developed system represents a step forward in DNA computing, combining for the first time DNA chemical processes with electronic input signals. Y1 - 2017 U6 - http://dx.doi.org/10.1002/elan.201600389 SN - 1521-4109 VL - 29 IS - 2 SP - 398 EP - 408 PB - Wiley-VCH CY - Weinheim ER -