AMMONIA-DEPENDENT SYNTHESIS AND METABOLIC CHANNELING OF CARBAMOYL-PHOSPHATE IN THE HYPERTHERMOPHILIC ARCHAEON PYROCOCCUS-FURIOSUS

The biosynthesis of carbamoyl phosphate (CP), a metabolic precursor of arginine and the pyrimidines was investigated in the hyperthermophili c archaeon Pyrococcus furiosus. The half-life of CP was found to be le ss than 2 s in the optimum temperature range of this organism (100-102 degrees C). The carbamoyl-phosphate synthase (CPSase) of P. furiosus uses ammonia as the nitrogen donor, and not glutamine like all micro-o rganisms investigated so far. The M(r) of the enzyme, which is devoid of regulatory properties, is 70000, at variance with that of known CPS ases. The possible significance of these findings with regard to hyper thermophilic nitrogen metabolism is discussed, Competition experiments with P. furiosus crude extracts indicated a marked preference of orni thine carbamoyltransferase (OTCase) for CP synthesized by CPSase rathe r than for CP added to the reaction mixture. In addition, the bisubstr ate analogue delta-N-phosphonoacetyl-L-ornithine inhibits the formatio n of citrulline from bicarbonate, ammonia, ATP and ornithine much less than its synthesis from ornithine and CP in the presence of free OTCa se. Such results suggest that, in vivo, CPSase and OTCase associate in a complex able to channel CP. Such a channelling may confer protectio n to CP, thus avoiding the accumulation of toxic amounts of cyanate ar ising from its decomposition as well as the waste of the two molecules of ATP required for its synthesis.