A gene encoding a putative GTP-specific phosphoenolpyruvate carboxykinase has been cloned and sequenced from the type I amitochondriate protist Giardia intestinalis. The deduced amino acid sequence is related most closely to homologs from hyperthermophilic archaebacteria and only more distantly to homologs from Eubacteria and Metazoa. Most enzymes of Giardia core metabolism, however, are related more closely to eubacterial and metazoan homologs. An archaebacterial relationship has been noted previously for the unusual acetyl-CoA synthetase (ADP-forming) of this organism. The results suggest that phosphoenolpyruvate carboxykinase and acetyl-CoA synthetase have been acquired from different sources than most enzymes of Giardia core metabolism.