GLUCOSE FERMENTATION TO ACETATE AND ALANINE IN RESTING CELL-SUSPENSIONS OF PYROCOCCUS-FURIOSUS - PROPOSAL OF A NOVEL GLYCOLYTIC PATHWAY BASED ON C-13 LABELING DATA AND ENZYME-ACTIVITIES

Suspensions of maltose-grown cells of the hyperthermophilic archaeon P yrococcus furiosus, when incubated at 90 degrees C with 35 mM [1-C-13] glucose or [3-C-13]glucose, consumed glucose at a rate of about 10 nmo l min(-1) (mg protein)(-1). Acetate (10 mM), alanine (3 mM), CO2 and H -2 were the fermentation products. The C-13-labelling pattern in alani ne and acetate were analyzed. With [1-C-13]glucose the methyl group of both alanine and acetate was labelled; with [3-C-13]glucose only the carboxyl group of alanine was labelled whereas acetate was unlabelled. Extracts of maltose-grown cells contained glucose isomerase (12.8 U m g(-1) 100 degrees C), ketohexokinase (0.23 U mg(-1), 100 degrees C), a nd fructose 1-phosphate aldolase (0.06 U mg(-1), 100 degrees C). Enzym es catalyzing the formation of fructose 1,6-bisphosphate from fructose 1-phosphate or fructose 6-phosphate could not be detected. As publish ed 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-Mey erhof pathway, and of 2-phosphoglycerate conversion - via pyruvate - t o acetate and alanine. Based on the enzyme activities in P. furiosus, the following pathway for glucose degradation to alanine and acetate i n cell suspensions is proposed which can explain the [C-13]glucose lab elling data: glucose --> fructose --> fructose 1-phosphate --> dihydro xyacetone phosphate + glyceraldehyde and further conversion of both tr ioses to alanine and acetate via pyruvate.