Phosphorylation of 3-O-methyl-d-glucose and catabolite repression in yeast

Abstract

The glucose analog, 3-0-methyl-~-glucose, inhibited growth of yeast on non-fermentable carbon sources. The sugar was phosphorylated by the yeast and also in vitro by a commercial preparation of yeast hexokinase. The chromatographic behaviour of the phosphorylated product was identical in both cases. This suggests that 3-0-methyl-D-glucose is phosphorylated to form 3-0-methyl-~-glucose 6-phosphate. The inhibition of the growth appears to be due to interference with the derepression of several enzymes necessary to grow on non-fermentable carbon sources. Spontaneous mutants whose growth was unaffected by 3-O-methyl-~-glucose were isolated. In these mutants there was no significant accumulation of the phosphorylated ester and the derepression of the enzymes tested was not affected by the glucose analog.Glucose causes inactivation and repression of several enzymes in yeast. These phenomena are known as catabolite inactivation [l] and catabolite repression [2]. The mechanisms underlying them are not well known although, in some particular cases, biochemical and genetic approaches have allowed the elaboration of a plausible hypothesis [3, 41. The use of some non-metabolizable glucose analogs like D-glucosamine and 2-deoxyglucose which are phosphorylated by yeast hexokinase has produced useful mutants for the study of catabolite repression [5, 61. It appeared of interest to evaluate the effects upon catabolite repression of a glucose analog not phosphorylated by hexokinase. We chose for this purpose 3-0-methyl-~-glucose (3 MeGlc) which has been shown to be a substrate for the glucose transport system in different organisms [7 -91 and appeared to be metabolically inert [ 101.This article shows that, contrary to current belief, 3 MeGlc is phosphorylated by yeast cells. This phosphorylation appears to be a prerequisite for the ability of the analog to interfere with the derepression of some enzymes subject to catabolite repression. MATERIALS AND METHODS Yeast strain andgrowth conditionsSaccharomyces cerevisiae F-11 a his1 ade can1 provided by Dr J. Conde (Sevilla, Spain) was used. The organisms were grown on liquid media with 1% yeast extract, 1% peptone and the adequate carbon source at a final concentration of 2%. Growth was followed turbidimetrically at 660 nm.

Galacturonic Acid Inhibits the Growth of Saccharomyces cerevisiae on Galactose, Xylose, and Arabinose

“…1 mol · g [dry weight] Ϫ1 ), which were in the same range as the concentrations of the hexose-phosphates that are intermediates of central yeast metabolism, such as glucose-6-phosphate or fructose-6-phosphate (7). Although not previously described for galacturonic acid 1-phosphate, the inhibitory effects of other phosphorylated compounds have been well documented (8,12,18,27,34). High levels of UDP-sugars can also have toxic effects (9), but the intracellular UDP-galacturonic acid concentration remained below the detection limit in this study.…”

Transcriptional Control of Gluconeogenesis in Aspergillus nidulans

“…In a creA204 background, AcuG-LacZ expression was elevated about twofold in the presence of glucose and sucrose ( Figure 4A). The effects of the nonmetabolizable analogs of glucose, 2-deoxy-d-glucose (2-DOG) and 3-O-methyl-d-glucose (3-MeGlc), able to bring about carbon catabolite repression in S. cerevisiae (Gancedo and Gancedo 1985), were also tested ( Figure Figure 3.-Analysis of expression of acuN and the effects of the acuN356 mutation. (A) Northern blot analysis of expression in a wild-type strain.…”

The Mechanism of Intracellular Acidification Induced by Glucose in Saccharomyces cerevisiae

“…As described previously (den Hollander et al, 1981 ; Nicolay et al, Caspani et al, 1985;Valle et al, 1986), the addition of 20 mM-glucose or other sugars which are degraded through glycolysis lowered pHin to about 6.4 (Table 1). Among the non-metabolizable sugars and sugar derivatives tested, those which are not efficiently phosphorylated (Sols et al, 1958;Gancedo & Gancedo, 1985;Siverio et al, 1986) such as xylose (150 mM), 3-0-methyl D-glucoside (20 mM) or 6-deoxy-~-glucose (100 mM) had little effect on pHin. However, 2-deoxy-~-glucose (20 mM), which is phosphorylated but not subsequently metabolized, caused a drop of pHin similar to that produced by glucose.…”