In vivo studies of pyridine nucleotide metabolism in Escherichia coli and Saccharomyces cerevisiae by carbon-13 NMR spectroscopy
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.
- Research Organization:
- Los Alamos National Lab., NM
- OSTI ID:
- 5819346
- Journal Information:
- J. Biol. Chem.; (United States), Vol. 259:4
- Country of Publication:
- United States
- Language:
- English
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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