Detection of modifications in the glucose metabolism induced by genetic mutations in Saccharomyces cerevisiae by C-13- and H-1-NMR spectroscopy

NMR spectroscopy may offer a suitable technique to characterize the glucose metabolism in response to genetic mutations in cells. The effects of vario us genetic modifications in Saccharomyces cerevisiae yeast were investigate d using C-13- and H-1-NMR spectroscopy associated with biochemical techniqu es. Cells were incubated with [1-C-13]glucose in order to study glucose con sumption and the formation of various end-products (ethanol, trehalose, gly cerol, glutamate and amino acids) as a function of time. Two types of genetic modifications were studied in S. cerevisiae. A genetic modification deleted the N-terminal part of the TFC7 protein which is the smallest subunit (tau55) of the TFIIIC transcription factor. One secondary effect of this mutation was a large deletion of mitochondrial DNA giving th e rho-phenotype. The other genetic modification corresponded to the disrupt ion of the HUF gene; the mutated cells were rho+ like the reference strain. Both mutations increase the glycolysis rate and glycerol synthesis and dec rease trehalose production. The most modified cells, which contain both TFC 7 deletion and HUF gene disruption, utilize glucose in the most extreme man ner as in these cells the largest production of the two glycolytic products (ethanol and glycerol) and the smallest trehalose formation occur. The HUF gene disruption serves as a positive modulator of glycolysis and respirati on. However, the TFC7 deletion, associated with the phenotype rho-, induces the most damage in the cellular function, dramatically altering the behavi our of the Krebs cycle. The cycle becomes blocked at the level of 2-oxoglut arate, detected by a characteristic pattern of the C-13-NMR glutamate spect ra. These NMR spectra corroborate the phenotypic data, the rho-phenotype co rresponding to deletions of mitochondria DNA which block all mitochondria p rotein synthesis and render the cells unable to derive energy from respirat ion. Moreover, as a consequence of the Krebs cycle blocking, alanine format ion is also observed.