A multinuclear magnetic resonance study of a cls11 mutant showing the Pet⁻ phenotype of Saccharomyces cerevisiae

Energetic and intermediary metabolism was studied in a Pet⁻ mutant of Saccharomyces cerevisiae with a calcium‐sensitive phenotype that shows an inability to grow when cultured in a medium containing non‐fermentable substrates. The perchloric acid extracts were prepared from suspensions of cls11 mutant and wild‐type cells incubated with [1,3‐¹³C]glycerol or [2‐¹³C]acetate, and analyzed by ³¹P, ¹³C and ¹H NMR. ³¹P‐ and ¹H‐NMR spectra showed significant differences between cls11 and wild‐type cells at the level of amino acids, the storage carbohydrate trehalose (higher in mutant cells), and sugar phosphates (higher in wild‐type cells). ¹³C‐NMR spectra revealed major differences in the steady‐state labelling of glutamate carbons. For incubations with [1,3‐¹³C]glycerol, we estimated from the relative ¹³C enrichment of glutamate carbons that acetyl‐CoA C2 is 43% C¹³ labelled in wild‐type and 10%¹³C labelled in mutant cells, respectively. For incubations with [2‐¹³C]acetate, we calculated that the ratio of the relative flux through the glyoxylate shunt versus oxidative reactions is 58% in wild‐type cells and 44% in the cls11 mutant cells. Again, a dilution of the relative enrichment of C2 of acetyl‐CoA was observed in the mutant cells (89%) compared to the wild‐type cells (97%). These results are discussed in terms of pleiotropic defects in non‐fermentable carbon metabolism in mutant cells.