In the yeast Saccharomyces cerevisiae, the enzyme gamma-glutamyl transpeptidase (gamma-GT; EC 2.3.2.2) is a glycoprotein that is bound to the vacuolar membrane. The kinetic parameters of GSH transport into isolated vacuoles were measured using intact vacuoles isolated from the wild-type yeast strain Sigma 1278b, under conditions of gamma-GT synthesis (nitrogen starvation) and repression (growth in the presence of ammonium ions). Vacuoles devoid of gamma-GT displayed a K(m) (app) of 18+/-2 mM and a V(max) (app) of 48.5+/-5 nmol of GSH/min per mg of protein. Vacuoles containing gamma-GT displayed practically the same K(m), but a higher V(max) (app) (150+/-12 nmol of GSH/min per mg of protein). Vacuoles prepared from a disruptant lacking gamma-GT showed no increase in V(max) (app) with nitrogen starvation. From a comparison of the transport data obtained for vacuoles isolated from various reference and mutant strains, it appears that the yeast cadmium factor 1 (YCF1) transport system accounts for approx. 70% of the GSH transport capacity of the vacuoles, the remaining 30% being due to a vacuolar (H(+)) ATPase-coupled system. The V(max) (app)-increasing effect of gamma-GT concerns only the YCF1 system. gamma-GT in the vacuolar membrane activates the Ycf1p transporter, either directly or indirectly. Moreover, GSH accumulating in the vacuolar space may exert a feedback effect on its own entry. Excretion of glutamate from radiolabelled GSH in isolated vacuoles containing gamma-GT was also measured. It is proposed that gamma-GT and a L-Cys-Gly dipeptidase catalyse the complete hydrolysis of GSH stored in the central vacuole of the yeast cell, prior to release of its constitutive amino acids L-glutamate, L-cysteine and glycine into the cytoplasm. Yeast appears to be a useful model for studying gamma-GT physiology and GSH metabolism.

In the yeast Saccharomyces cerevisiae, the enzyme γ-glutamyl transpeptidase (γ-GT; EC 2.3.2.2) is a glycoprotein that is bound to the vacuolar membrane. The kinetic parameters of GSH transport into isolated vacuoles were measured using intact vacuoles isolated from the wild-type yeast strain Σ1278b, under conditions of γ-GT synthesis (nitrogen starvation) and repression (growth in the presence of ammonium ions). Vacuoles devoid of γ-GT displayed a Km (app) of 18±2mM and a Vmax (app) of 48.5±5nmol of GSH/min per mg of protein. Vacuoles containing γ-GT displayed practically the same Km, but a higher Vmax (app) (150±12nmol of GSH/min per mg of protein). Vacuoles prepared from a disruptant lacking γ-GT showed no increase in Vmax (app) with nitrogen starvation. From a comparison of the transport data obtained for vacuoles isolated from various reference and mutant strains, it appears that the yeast cadmium factor 1 (YCF1) transport system accounts for approx. 70% of the GSH transport capacity of the vacuoles, the remaining 30% being due to a vacuolar (H+) ATPase-coupled system. The Vmax (app)-increasing effect of γ-GT concerns only the YCF1 system. γ-GT in the vacuolar membrane activates the Ycf1p transporter, either directly or indirectly. Moreover, GSH accumulating in the vacuolar space may exert a feedback effect on its own entry. Excretion of glutamate from radiolabelled GSH in isolated vacuoles containing γ-GT was also measured. It is proposed that γ-GT and a l-Cys-Gly dipeptidase catalyse the complete hydrolysis of GSH stored in the central vacuole of the yeast cell, prior to release of its constitutive amino acids l-glutamate, l-cysteine and glycine into the cytoplasm. Yeast appears to be a useful model for studying γ-GT physiology and GSH metabolism.

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The Luedeking-Piret model is an empirical relationship which is very widely used in cell cultures to evaluate specific production rates of some products (metabolites or others). It constitutes a very common method of calculation as much in fundamental as in applied research and especially for designing and optimizing industrial processes in very varied fields. However, this model appears to be frequently deficient and has to be greatly adapted, practically, one might say, for each individual case. Obviously, this is a very great drawback, requiring a great deal of time spent on it and one that greatly lessens the 'universality' of the model. This work reveals that it is possible to give the initial Luedeking-Piret model a much more general scope. The used method revealed metabolic switches that have never been suspected until now. Confirmation of the method would certainly give a precious general tool both to optimize production processes and to increase understanding of some physiological states of cells in chemostat.