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Estimation of intracellular sugar phosphate concentrations in Saccharomyces cerevisiae using 31 P nuclear magnetic resonance spectroscopy
Author(s) -
Shanks Jacqueline Vanni,
Bailey James E.
Publication year - 1988
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.260320907
Subject(s) - sugar phosphates , fructose , intracellular , phosphate , chemistry , nuclear magnetic resonance spectroscopy , saccharomyces cerevisiae , biochemistry , sugar , polyphosphate , glucose 6 phosphate , intracellular ph , in vivo , yeast , biophysics , biology , stereochemistry , enzyme , microbiology and biotechnology
A systematic procedure has been formulated for estimating the relative intracellular concentrations of sugar phosphates in Saccharomyces cerevisiae based upon 31 P nuclear magnetic resonance (NMR) measurements. The sugar phosphate region of the 31 P NMR spectrum is first decomposed by computer analysis, and the decomposition consistency and identification of individual sugar phosphate resonances are established based on in vitro chemical shift calibrations determined in separate experiments. Numerous evaluations of intracellular S. cerevisiae compositions for different strains and different cell environments provide the basis for in vivo correlations of inorganic phosphate chemical shift with the chemical shifts of 3‐phosphoglycerate, beta;‐fructose 1,6‐diphosphate, fructose 6‐phosphate, and glucose 6 phosphate. Relative intracellular sugar phosphate concentrations are obtained by correcting peak areas for partial saturation during transient in vivo experiments. In vivo concentrations estimated by this method agree well with estimates for similar systems based on other techniques. This approach does not require costly la belled compounds, and has the advantage that other important metabolic state variables such‐as internal and external pH and intracellular levels of phosphate, ATP, ADP, NAD(H), and polyphosphate may be determined from the same 31 P spectrum. Extension of this strategy to other cellular systems should be straightforward.