Comparison of wild-type and Reg 1 mutant saccharomyces cerevisiae metabolic levels during glucose and galactose metabolism using 31P NMR.
The reg 1 mutation will allow the expression of a cloned gene on a plasmid under the control of a GAL promoter in the presence of glucose. The metabolism of wild-type and reg l mutant strains was examined by in vivo (31)P nuclear magnetic resonance (NMR) spectroscopy. Transient profiles of glucose 6-phosphate, fructose 6-phosphate, fructose 1, 6-diphosphate, and 3-phosphoglycerate indicated that glucose was processed differently for the reg 1 strain despite similar cytoplasrnic pH values and ATP levels. Intracellular phosphate became depleted in the transition to quasi-steady state and limited glycolysis in the reg 1 strain. The glucose uptake step or hexokinase step appears to be altered in the reg 1 strain. The reg 1 strain utilized galactose faster than the wild-type strain under the conditions used for NMR analysis. These results are consistent with the hypothesis that the REG 1 product operates early in the regulatory circuitry for glucose repression. This study illustrates the usefulness of transient information provided by NMR in understanding changes in the metabolism of genetically manipulated organisms.
Shanks
J V
JV
Department of Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.
Bailey
J E
JE