Re-investigation of glucose metabolism in Fibrobacter succinogenes, using NMR spectroscopy and enzymatic assays. : Evidence for pentose phosphates phosphoketolase and pyruvate formate lyase activities

Abstract

The glucose metabolism of Fibrobacter succinogenes S85 was studied in detail; key intermediates and alternative pathways were evidenced by NMR and/or enzymatic assays. A high phosphoketolase activity was detected in four strains of Fibrobacter under strictly anaerobic conditions, with ribose-5-phosphate as substrate, no activity was evidenced with fructose-6-phosphate. This is the first report of a pentose phosphates phosphoketolase in bacteria unable to use pentoses. In contrast, the Entner-Doudoroff pathway and the oxidative branch of the pentose phosphate pathway could not be evidenced. Incubation of living cells of F. succinogenes with Na 2 13CO 3 confirmed the incorporation of 13CO 2 in the carboxylic group of succinate. The presence of fumarase was evidenced by in vivo 13C-NMR using 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO); the enzyme showed a high reversibility under physiological conditions. The production of formate from glucose catabolism was evidenced by enzymatic assay and by NMR and a pyruvate formate lyase activity was detected using strictly anaerobic conditions.