Real-time detection of cofactor availability in genetically modified living Saccharomyces cerevisiae cells - Simultaneous probing of different geno- and phenotypes

Natalie Kostesha, Arto Heiskanen, C. Spegel, B Hahn-Hagerdal, M.F. Gorwa-Grauslund, Jenny Emnéus

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    This work describes a mediated amperometric method for simultaneous real-time probing of the NAD(P)H availability in two different phenotypes, fermentative and respiratory, of the phosphoglucose isomerase deletion mutant strain of S. cerevisiae. EBY44 [ENY.WA-1A pgi1-1D::URA3], and its parental strain, ENY.WA-1A. The developed method is based on multichannel detection using microelectrode arrays. Its versatility was demonstrated by using four microelectrode arrays for simultaneously monitoring the NAD(P)H availability of both geno- and phenotypes under the influence of two different carbon sources, glucose and fructose, as well as the cytosolic and mitochondrial inhibitor and uncoupler, dicoumarol. The obtained results indicate that the method is capable of accurately and reproducibly (overall relative standard error of mean 3.2%) mapping the real-time responses of the cells with different genotype-phenotype combinations. The ENY.WA cells showed the same response to glucose and fructose when dicoumarol was used; fermentative cells indicated the presence of cytosolic inhibition and respiratory cells a net effect of mitochondrial uncoupling. EBY44 cells showed cytosolic inhibition with the exception of respiratory cells when fructose was used as carbon source. (C) 2009 Elsevier B.V. All rights reserved.
    Original languageEnglish
    JournalBioelectrochemistry
    Volume76
    Issue number1-2
    Pages (from-to)180-188
    ISSN1567-5394
    DOIs
    Publication statusPublished - 2009

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