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Amperometric response from the glycolytic versus the pentose phosphate pathway in Saccharomyces cerevisiae cells

Spegel, Christer LU ; Heiskanen, Arto LU ; Kostesha, Natalie LU ; Johanson, Ted LU ; Gorwa-Grauslund, Marie-Francoise LU ; Koudelka-Hep, M; Emnéus, Jenny LU and Ruzgas, Tautgirdas LU (2007) In Analytical Chemistry 79(23). p.8919-8926
Abstract
The two main metabolic pathways involved in sugar metabolism, i.e., the pentose phosphate pathway (PPP) and the glycolytic pathway (GP), were amperometrically monitored using a double-mediator system composed of menadione and ferricyanide. With the use of the Saccharomyces cerevisiae deletion mutant, EBY44, lacking the gene encoding for the branch point enzyme phosphoglucose isomerize, selective amperometric monitoring of the PPP, mainly producing NADPH, and the GP, mainly producing NADH, could be achieved. It was found that the bioelectrocatalytic current was primarily originating from NADPH. This conclusion was supported by metabolite flux analysis, confirming that, in the presence of menadione, the cells increase the rate of... (More)
The two main metabolic pathways involved in sugar metabolism, i.e., the pentose phosphate pathway (PPP) and the glycolytic pathway (GP), were amperometrically monitored using a double-mediator system composed of menadione and ferricyanide. With the use of the Saccharomyces cerevisiae deletion mutant, EBY44, lacking the gene encoding for the branch point enzyme phosphoglucose isomerize, selective amperometric monitoring of the PPP, mainly producing NADPH, and the GP, mainly producing NADH, could be achieved. It was found that the bioelectrocatalytic current was primarily originating from NADPH. This conclusion was supported by metabolite flux analysis, confirming that, in the presence of menadione, the cells increase the rate of NADPH-producing reactions although these processes might be detrimental to cell survival. The higher rate of in vivo NADPH-dependent menadione reduction can be ascribed to the fact that the intracellular NADPH/NADP+ ratio is much higher than NADH/NAD+ as well as that the former ratio is more tightly controlled. This tight control over the cofactor ratios is lost upon cell disintegration as observed from spectrophotometric assays using crude cell extract, and amperometric investigations of permeabilized cells indicate a higher rate of NADH- than NADPH-dependent menadione reduction. These in vitro experiments show a higher activity of NADH-dependent than NADPH-dependent menadione-reducing dehydrogenases in S. cerevisiae cells. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Analytical Chemistry
volume
79
issue
23
pages
8919 - 8926
publisher
The American Chemical Society
external identifiers
  • wos:000251311900018
  • scopus:36849086459
ISSN
1520-6882
DOI
10.1021/ac0710679
language
English
LU publication?
yes
id
f3461b16-8fea-479c-b1bb-d530c4d1d473 (old id 935709)
date added to LUP
2008-01-17 17:03:38
date last changed
2017-11-12 03:18:35
@article{f3461b16-8fea-479c-b1bb-d530c4d1d473,
  abstract     = {The two main metabolic pathways involved in sugar metabolism, i.e., the pentose phosphate pathway (PPP) and the glycolytic pathway (GP), were amperometrically monitored using a double-mediator system composed of menadione and ferricyanide. With the use of the Saccharomyces cerevisiae deletion mutant, EBY44, lacking the gene encoding for the branch point enzyme phosphoglucose isomerize, selective amperometric monitoring of the PPP, mainly producing NADPH, and the GP, mainly producing NADH, could be achieved. It was found that the bioelectrocatalytic current was primarily originating from NADPH. This conclusion was supported by metabolite flux analysis, confirming that, in the presence of menadione, the cells increase the rate of NADPH-producing reactions although these processes might be detrimental to cell survival. The higher rate of in vivo NADPH-dependent menadione reduction can be ascribed to the fact that the intracellular NADPH/NADP+ ratio is much higher than NADH/NAD+ as well as that the former ratio is more tightly controlled. This tight control over the cofactor ratios is lost upon cell disintegration as observed from spectrophotometric assays using crude cell extract, and amperometric investigations of permeabilized cells indicate a higher rate of NADH- than NADPH-dependent menadione reduction. These in vitro experiments show a higher activity of NADH-dependent than NADPH-dependent menadione-reducing dehydrogenases in S. cerevisiae cells.},
  author       = {Spegel, Christer and Heiskanen, Arto and Kostesha, Natalie and Johanson, Ted and Gorwa-Grauslund, Marie-Francoise and Koudelka-Hep, M and Emnéus, Jenny and Ruzgas, Tautgirdas},
  issn         = {1520-6882},
  language     = {eng},
  number       = {23},
  pages        = {8919--8926},
  publisher    = {The American Chemical Society},
  series       = {Analytical Chemistry},
  title        = {Amperometric response from the glycolytic versus the pentose phosphate pathway in Saccharomyces cerevisiae cells},
  url          = {http://dx.doi.org/10.1021/ac0710679},
  volume       = {79},
  year         = {2007},
}