Glucose fermentation to acetate and alanine in resting cell suspensions ofPyrococcus furiosus: Proposal of a novel glycolytic pathway based on¹³C labelling data and enzyme activities

Suspensions of maltose-grown cells of the hyperthermophilic archaeon Pyrococcus furiosus , when incubated at 90°C with 35 mM [1- 13 C]glucose or [3- 13 C]glucose, consumed glucose at a rate of about 10 nmol min −1 (mg protein) −1 . Acetate (10 mM), alanine (3 mM), CO 2 and H 2 were the fermentation products. The 13 C-labelling pattern in alamine and acetate were analyzed. With [1- 13 C]glucose the methyl group of both alanine and acetate was labelled; with [3- 13 C]glucose only the carboxyl group of alanine was labelled whereas acetate was unlabelled. Extracts of maltose-grown cells contained glucose isomerase (12.8 U mg −1 , 100°C), ketohexokinase (0.23 U mg −1 , 100°C), and fructose 1-phosphate aldolase (0.06 U mg −1, 100°C). Enzymes catalyzing the formation of fructose 1,6-bisphosphate from fructose 1-phosphate or fructose 6-phosphate could not be detected. As publihed previously by our group and other authors P. furiosus also contains enzymes of glyceraldehyde conversion to 2-phosphoglycerate according to a non-phosphorylated Entner-Doudoroff pathway, of dihydroxyacetone phosphate conversion to 2-phosphoglycerate according to the Embden-Meyerhof pathway, and of 2-phosphoglycerate conversion - via pyruvate - to acetate and alanine. Based on the enzyme activities in P. furiosus , the following pathway for glucose degradation to alanine and acetate in cell suspensions is proposed which can explain the [ 13 C]glucose labelling data: glucose→ fructose → fructose 1- phosphate → dihydroxyacetone phosphate + glyceraldehyde and further conversion of both trioses to alanine and acetate via pyruvate.