The effect of increased phosphoglucose isomerase on glucose metabolism in Saccharomyces cerevisiae

Abstract

Comparison of microbial strains with normal and high content of single enzymes is coming into use for metabolic analysis and in vivo assessment of enzyme function. We present an example for phosphoglucose isomerase and glucose metabolism in the yeast Saccharomyces cerevisiae. We use cell suspensions in conditions of inhibited protein synthesis and respiration, with low assimilation, rapid and linear glucose utilization, fermentation almost quantitative, and high enough cell density for direct preparation of extracts for metabolite analysis. The mass action ratio and fitting of fructose-6-P and glucose-6-P concentrations and kinetic parameters of the enzyme are not inconsistent with near equilibrium of the reaction in the wild-type strain and small if any change in the high level strain. However, this conclusion would require that the Vmax values underestimate the activity in the cell. On the other hand, the specific activities of glucose-6-P and fructose-1,6-P2 during metabolism of [2-3H]glucose are quite high which, together with knowledge of tritium exchange and isotope effects for the reaction in vitro, would point to the reaction in the wild-type strain being far from equilibrium; the specific activities are lower in the high level strain, indicating that extra enzyme is functional. One way to reconcile the latter results would be for tritium exchange to be considerably lower in vivo than known in vitro.