Abstract

Glycerol is highly abundant in nature and serve as carbon source for many organisms. Also, several Archaea have the genetic capacity to grow on glycerol but its degradation has so far only been studied Haloferax volcanii. Herein, the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius was shown to grow with glycerol as sole carbon and energy source. After uptake likely involving facilitated diffusion, glycerol is degraded via phosphorylation to glycerol-3-phosphate followed by oxidation to dihydroxyacetone phosphate (DHAP) catalyzed by glycerol kinase (GK) by an unusual quinone reducing FAD-dependent glycerol-3-phosphate dehydrogenase (G3PDH), respectively. The S. acidocaldarius genome harbors two paralogous copies of each GK and G3PDH. However, only one of these GK-G3PDH couples (Saci_2031-2033) is highly upregulated on glycerol. Deletion of the saci_2033 gene encoding GK abolished growth on glycerol and GK activity in crude extracts. In contrast, deletion of the second GK gene (saci_1117) had only minor effects indicating that only one of the two GK-G3PDH couples is essential. Biochemical characterization revealed that both isoenzymes of each, GK and G3PDH, were functionally similar. Whereas the GKs showed high similarity to known enzymes from Bacteria and Eukaryotes, the G3PDHs represent unusual homologues of the bacterial GlpA subunit of the GlpABC complex with remarkable C-terminal sequence differences and a novel type of membrane anchoring via a CoxG-like protein (Saci_2031). Further sequence analyzes discovered a higher versatility of G3PDHs in Archaea with respect to interacting proteins, electron transfer, and membrane anchoring likely reflecting tailored evolutionary solutions to meet different requirements caused by life styles and electron acceptors.