Kinetics of Sugar Transport and Phosphorylation Influence Glucose and Fructose Cometabolism by Zymomonas mobilis

Abstract

The competitive inhibition of fructokinase by glucose has been proposed as the mechanism by which Zymomonas mobilis preferentially consumes glucose from mixtures of glucose and fructose and accumulates fructose when growing on sucrose. In this study, incorporation of radioactive fructose into biomass was used as a measure of fructose catabolism. It was determined that the rate of fructose incorporation by Z. mobilis CP4 was somewhat lower in the presence of an equimolar concentration of glucose but that the inhibition of fructokinase by glucose was not nearly as severe in vivo as was predicted from in vitro studies. Interestingly, addition of glucose to a culture of Z. mobilis CP4-M2, a glucokinaseless mutant, resulted in an immediate and nearly complete inhibition of fructose incorporation. Furthermore, addition of nonmetabolizeable glucose analogs had a similar effect on fructose catabolism by the wild-type Z. mobilis CP4, and fructose uptake by Z. mobilis CP4-M2 was shown to be severely inhibited by equimolar amounts of glucose. These results suggest that competition for fructose transport plays an important role in preferential catabolism of glucose from sugar mixtures. Indeed, the apparent K(infm) values for sugar uptake by Z. mobilis CP4 were approximately 200 mM for fructose and 13 mM for glucose. Other experiments supported the conclusion that a single facilitated diffusion transport system, encoded by the glf gene, is solely responsible for the uptake of both glucose and fructose. The results are discussed with regard to the hypothesis that the kinetics of sugar transport and phosphorylation allow the preferential consumption of glucose and accumulation of fructose, making the fructose available for the enzyme glucose-fructose oxidoreductase, which forms sorbitol, an important osmoprotectant for Z. mobilis when growing in the presence of high sugar concentrations.