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Title: In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy

Abstract

Pyridine nucleotide metabolism has been studied in vivo in a prokaryotic (Escherichia coli) and a eukaryotic (Saccharomyces cerevisiae) system cultured in a medium containing carbon-13-labeled nicotinic acid, followed by NMR detection of the labeled organisms. Chemical exchange between oxidized and reduced nucleotides is found to be sufficiently slow on the NMR time scale to permit the observation of separate resonances corresponding to each redox state. The possibility of significant exchange broadening of reduced pyridine nucleotide resonances under some conditions was further evaluated based on comparative NMR studies utilizing organisms cultured in the presence of either (2-/sup 13/C)nicotinate or (5-/sup 13/C)nicotinate. Based on these experiments, it was concluded that broadening as a consequence of intermediate exchange is not significant. Although it was initially anticipated that the carbon-13 resonances arising from the di- and triphosphopyridine nucleotide pools could not be distinguished, the absence of observable resonances corresponding to reduced nucleotides in oxygenated yeast and E. coli cells suggests that the NMR method is fairly specific for determining the redox status of the diphosphopyridine nucleotide pool. Studies of the effects of a variety of perturbations including variation of the oxygen supply, addition of ethanol, and addition of the oxidative phosphorylation uncoupler dinitrophenol havemore » been carried out. Dramatic differences in the response of the catabolic reduction charge, CRC = (NADH)/(NADH) + (NAD/sup +/), between the yeast and E. coli cells are observed. The CRC values for the yeast undergo large changes in response to these perturbations which are not observed for the bacterial cells. 52 references, 9 figures, 2 tables.« less

Authors:
;
Publication Date:
Research Org.:
Los Alamos National Lab., NM
OSTI Identifier:
5819346
Resource Type:
Journal Article
Journal Name:
J. Biol. Chem.; (United States)
Additional Journal Information:
Journal Volume: 259:4
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; PYRIDINES; METABOLISM; CARBON 13; ESCHERICHIA COLI; ETHANOL; EXPERIMENTAL DATA; IN VIVO; NAD; NADH2; NICOTINIC ACID; NITROPHENOL; NUCLEAR MAGNETIC RESONANCE; OXYGEN; SACCHAROMYCES CEREVISIAE; ALCOHOLS; AROMATICS; AZINES; BACTERIA; CARBON ISOTOPES; CARBOXYLIC ACIDS; COENZYMES; DATA; ELEMENTS; EVEN-ODD NUCLEI; FUNGI; HETEROCYCLIC ACIDS; HETEROCYCLIC COMPOUNDS; HYDROXY COMPOUNDS; INFORMATION; ISOTOPES; LIGHT NUCLEI; MAGNETIC RESONANCE; MICROORGANISMS; NITRO COMPOUNDS; NONMETALS; NUCLEI; NUCLEOTIDES; NUMERICAL DATA; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANIC NITROGEN COMPOUNDS; PHENOLS; PLANTS; RESONANCE; SACCHAROMYCES; STABLE ISOTOPES; VITAMIN B GROUP; VITAMINS; YEASTS; 550501* - Metabolism- Tracer Techniques

Citation Formats

Unkefer, C J, and London, R E. In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy. United States: N. p., 1984. Web.
Unkefer, C J, & London, R E. In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy. United States.
Unkefer, C J, and London, R E. Sat . "In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy". United States.
@article{osti_5819346,
title = {In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy},
author = {Unkefer, C J and London, R E},
abstractNote = {Pyridine nucleotide metabolism has been studied in vivo in a prokaryotic (Escherichia coli) and a eukaryotic (Saccharomyces cerevisiae) system cultured in a medium containing carbon-13-labeled nicotinic acid, followed by NMR detection of the labeled organisms. Chemical exchange between oxidized and reduced nucleotides is found to be sufficiently slow on the NMR time scale to permit the observation of separate resonances corresponding to each redox state. The possibility of significant exchange broadening of reduced pyridine nucleotide resonances under some conditions was further evaluated based on comparative NMR studies utilizing organisms cultured in the presence of either (2-/sup 13/C)nicotinate or (5-/sup 13/C)nicotinate. Based on these experiments, it was concluded that broadening as a consequence of intermediate exchange is not significant. Although it was initially anticipated that the carbon-13 resonances arising from the di- and triphosphopyridine nucleotide pools could not be distinguished, the absence of observable resonances corresponding to reduced nucleotides in oxygenated yeast and E. coli cells suggests that the NMR method is fairly specific for determining the redox status of the diphosphopyridine nucleotide pool. Studies of the effects of a variety of perturbations including variation of the oxygen supply, addition of ethanol, and addition of the oxidative phosphorylation uncoupler dinitrophenol have been carried out. Dramatic differences in the response of the catabolic reduction charge, CRC = (NADH)/(NADH) + (NAD/sup +/), between the yeast and E. coli cells are observed. The CRC values for the yeast undergo large changes in response to these perturbations which are not observed for the bacterial cells. 52 references, 9 figures, 2 tables.},
doi = {},
url = {https://www.osti.gov/biblio/5819346}, journal = {J. Biol. Chem.; (United States)},
number = ,
volume = 259:4,
place = {United States},
year = {1984},
month = {2}
}