Owing to the absence of the pentose phosphate pathway, the degradation pathway for the ribose moieties of nucleosides is unknown in Archaea. Here, in the archaeon Thermococcus kodakarensis, we identified a metabolic network that links the pentose moieties of nucleosides or nucleotides to central carbon metabolism. The network consists of three nucleoside phosphorylases, an ADP-dependent ribose-1-phosphate kinase and two enzymes of a previously identified NMP degradation pathway, ribose-1,5-bisphosphate isomerase and type III ribulose-1,5-bisphosphate carboxylase/oxygenase. Ribose 1,5-bisphosphate and ribulose 1,5-bisphosphate are intermediates of this pathway, which is thus designated the pentose bisphosphate pathway.

dAMP phosphorylase (AMPpase), ribose-1,5-bisphosphate (R15P) isomerase, and type III ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) have been proposed to constitute a novel pathway involved in AMP metabolism in the Archaea. Here we performed a biochemical examination of AMPpase and R15P isomerase from Thermococcus kodakarensis. R15P isomerase was specific for the ␣-anomer of R15P and did not recognize other sugar compounds. We observed that activity was extremely low with the substrate R15P alone but was dramatically activated in the presence of AMP. Using AMP-activated R15P isomerase, we reevaluated the substrate specificity of AMPpase. AMPpase exhibited phosphorylase activity toward CMP and UMP in addition to AMP. The [S]-v plot (plot of velocity versus substrate concentration) of the enzyme toward AMP was sigmoidal, with an increase in activity observed at concentrations higher than approximately 3 mM. The behavior of the two enzymes toward AMP indicates that the pathway is intrinsically designed to prevent excess degradation of intracellular AMP. We further examined the formation of 3-phosphoglycerate from AMP, CMP, and UMP in T. kodakarensis cell extracts. 3-Phosphoglycerate generation was observed from AMP alone, and from CMP or UMP in the presence of dAMP, which also activates R15P isomerase. 3-Phosphoglycerate was not formed when 2-carboxyarabinitol 1,5-bisphosphate, a Rubisco inhibitor, was added. The results strongly suggest that these enzymes are actually involved in the conversion of nucleoside monophosphates to 3-phosphoglycerate in T. kodakarensis.A rchaea comprise the third domain of life and exhibit unique metabolic features that are not found in bacteria and eukaryotes. The metabolic enzymes and pathways utilized for glycolysis and pentose biosynthesis in many archaea differ from the classical Embden-Meyerhof (EM)/Entner-Doudoroff (ED) pathways (24,27,33,35) and the pentose phosphate pathway (11,19,28), respectively. A previous study of the hyperthermophilic archaeon Thermococcus kodakarensis KOD1 (3, 9) suggested the presence of a novel pathway involved in nucleic acid metabolism (25). The pathway consists of a type III ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and two novel enzymes, AMP phosphorylase (AMPpase) and ribose-1,5-bisphosphate (R15P) isomerase. In the first reaction of this pathway, catalyzed by AMPpase, the adenine base of AMP is released and is replaced by a phosphate group to generate R15P. In the following R15P isomerase reaction, R15P is converted to ribulose 1,5-bisphosphate (RuBP). In the final reaction, Rubisco catalyzes the conversion of RuBP, CO 2 , and H 2 O to 2 molecules of 3-phosphoglycerate (3-PGA), which is an intermediate of central sugar metabolism. Genome sequences indicate that this pathway is distributed broadly among the Archaea, including all members of the Thermococcales, Archaeoglobales, Methanomicrobiales, and Methanosarcinales. The pathway is also found in several members of the Halobacteriales, Methanococcales, Desulfurococcales...

Background: Ribose-1,5-bisphosphate isomerase (R15Pi) converts ribose 1,5-bisphosphate into ribulose 1,5-bisphosphate in a novel AMP metabolic pathway. Results: Crystal structures of reaction-ready and -completed states are determined. Conclusion:R15Pi undergoes an open-closed conformational change upon substrate binding, and the reaction proceeds via a cis-phosphoenolate intermediate. Significance:The mechanism of ribose isomerization revealed in this study could be applied on other 1-phosphorylated ribose isomerases.