EVIDENCE FOR THE OPERATION OF A NOVEL EMBDEN-MEYERHOF PATHWAY THAT INVOLVES ADP-DEPENDENT KINASES DURING SUGAR FERMENTATION BY PYROCOCCUS-FURIOSUS

The main pathway for the fermentation of maltose or cellobiose by the hyperthermophile Pyrococcus furiosus was investigated by in vivo NMR a nd by enzyme measurements. Addition of [1-C-13]glucose to cell suspens ions resulted in the formation of C-2-labeled acetate and C-3- labeled alanine. No label was recovered in CO2 or HCO3-. In the presence of [ 3-C-13]glucose, the label ended up in the C-1 atom of alanine and in H CO3- and CO2. These labeling patterns indicate that glucose is convert ed along an Embden-Meyerhof pathway, and they disagree with the previo usly proposed nonphosphorylated Entner-Doudoroff pathway (pyroglycolys is). The NMR data were supported by enzyme measurements. Hexokinase (8 .7 units/mg), phosphoglucose isomerase (6.8 units/mg), phosphofructoki nase (0.81 unit/mg), and aldolase (0.26 unit/mg) were present in cell- free extracts (specific activities at 90 degrees C). Remarkably, the t wo kinases required ADP as the phosphoryl group donor instead of ATP. No activity was found with pyrophosphate. These are the first descript ions of ADP-dependent (AMP-forming) kinases to date. Since P. furiosus is a phylogenetically ancient organism, these enzymes may represent a n ancestral kind of metabolism.