Reference "Analysis of the compartmentation of glycolytic intermediates, nucleotides, sugars, organic acids, amino acids, and sugar alcohols in potato tubers using a nonaqueous fractionation method"

Reference ID

54965

Title

Analysis of the compartmentation of glycolytic intermediates, nucleotides, sugars, organic acids, amino acids, and sugar alcohols in potato tubers using a nonaqueous fractionation method

Source

Plant Physiol, 2001, vol. 127, pp. 685-700

Authors (6)

Farre-E-M	Tiessen-A
Roessner-U	Geigenberger-P
Trethewey-R-N	Willmitzer-L

Abstract

The compartmentation of metabolism in heterotrophic plant tissues is poorly
understood due to the lack of data on metabolite distributions and fluxes
between subcellular organelles. The main reason for this is the lack of suitable
experimental methods with which intracellular metabolism can be measured. Here,
we describe a nonaqueous fractionation method that allows the subcellular
distributions of metabolites in developing potato (Solanum tuberosum L. cv
Desiree) tubers to be calculated. In addition, we have coupled this
fractionation method to a recently described gas chromatography-mass
spectrometry procedure that allows the measurement of a wide range of small
metabolites. To calculate the subcellular metabolite concentrations, we have
analyzed organelle volumes in growing potato tubers using electron microscopy.
The relative volume distributions in tubers are very similar to the ones for
source leaves. More than 60% of most sugars, sugar alcohols, organic acids, and
amino acids were found in the vacuole, although the concentrations of these
metabolites is often higher in the cytosol. Significant amounts of the
substrates for starch biosynthesis, hexose phosphates, and ATP were found in the
plastid. However, pyrophosphate was located almost exclusively in the cytosol.
Calculation of the mass action ratios of sucrose synthase, UDP-glucose
pyrophosphorylase, phosphoglucosisomerase, and phosphoglucomutase indicate that
these enzymes are close to equilibrium in developing potato tubers. However, due
to the low plastidic pyrophosphate concentration, the reaction catalyzed by ADP-
glucose pyrophosphorylase was estimated to be far removed from equilibrium.