| 1. |
- Heiskanen, Arto, et al.
(författare)
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Bioelectrochemical probing of intracellular redox processes in living yeast cells-application of redox polymer wiring in a microfluidic environment
- 2013
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Ingår i: Analytical and Bioanalytical Chemistry. - : Springer Science and Business Media LLC. - 1618-2642 .- 1618-2650. ; 405:11, s. 3847-3858
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Tidskriftsartikel (refereegranskat)abstract
- Conventionally, microbial bioelectrochemical assays have been conducted using immobilized cells on an electrode that is placed in an electrochemical batch cell. In this paper, we describe a developed microfluidic platform with integrated microelectrode arrays for automated bioelectrochemical assays utilizing a new double mediator system to map redox metabolism and screen for genetic modifications in Saccharomyces cerevisiae cells. The function of this new double mediator system based on menadione and osmium redox polymer (PVI-Os) is demonstrated. "Wiring" of S. cerevisiae cells using PVI-Os shows a significant improvement of bioelectrochemical monitoring in a microfluidic environment and functions as an effective immobilization matrix for cells that are not strongly adherent. The function of the developed microfluidic platform is demonstrated using two strains of S. cerevisiae, ENY. WA and its deletion mutant EBY44, which lacks the enzyme phosphoglucose isomerase. The cellular responses to introduced glucose and fructose were recorded for the two S. cerevisiae strains, and the obtained results are compared with previously published work when using an electrochemical batch cell, indicating that microfluidic bioelectrochemical assays employing the menadione-PVI-Os double mediator system provides an effective means to conduct automated microbial assays.
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| 2. |
- Kostesha, Natalie, et al.
(författare)
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Probing the redox metabolism in the strictly anaerobic, extremely thermophilic, hydrogen-producing Caldicellulosiruptor saccharolyticus using amperometry
- 2011
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Ingår i: Extremophiles. - : Springer Science and Business Media LLC. - 1433-4909 .- 1431-0651. ; 15:1, s. 77-87
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Tidskriftsartikel (refereegranskat)abstract
- Changes in the redox metabolism in the anaerobic, extremely thermophilic, hydrogen-forming bacterium Caldicellulosiruptor saccharolyticus were probed for the first time in vivo using mediated amperometry with ferricyanide as a thermotolerant external mediator. Clear differences in the intracellular electron flow were observed when cells were supplied with different carbon sources. A higher electrochemical response was detected when cells were supplied with xylose than with sucrose or glucose. Moreover, using the mediated electrochemical method, it was possible to detect differences in the electron flow between cells harvested in the exponential and stationary growth phases. The electron flow of C. saccharolyticus was dependent on the NADH- and reduced ferredoxin generation flux and the competitive behavior of cytosolic and membrane-associated oxidoreductases. Sodium oxamate was used to inhibit the NADH-dependent lactate dehydrogenase, upon which more NADH was directed to membrane-associated enzymes for ferricyanide reduction, leading to a higher electrochemical signal. The method is noninvasive and the results presented here demonstrate that this method can be used to accurately detect changes in the intracellular electron flow and to probe redox enzyme properties of a strictly anaerobic thermophile in vivo.
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