title: PPi-dependent phosphofructokinase from Thermoproteus tenax, an archaeal descendant of an ancient line in phosphofructokinase evolution.
abstract: Flux into the glycolytic pathway of most cells is controlled via allosteric regulation of the irreversible, committing step catalyzed by ATP-dependent phosphofructokinase (PFK) (ATP-PFK; EC 2.7.1.11), the key enzyme of glycolysis. In some organisms, the step is catalyzed by PPi-dependent PFK (PPi-PFK; EC 2.7.1.90), which uses PPi instead of ATP as the phosphoryl donor, conserving ATP and rendering the reaction reversible under physiological conditions. We have determined the enzymic properties of PPi-PFK from the anaerobic, hyperthermophilic archaeon Thermoproteus tenax, purified the enzyme to homogeneity, and sequenced the gene. The approximately 100-kDa PPi-PFK from T. tenax consists of 37-kDa subunits; is not regulated by classical effectors of ATP-PFKs such as ATP, ADP, fructose 2,6-bisphosphate, or metabolic intermediates; and shares 20 to 50% sequence identity with known PFK enzymes. Phylogenetic analyses of biochemically characterized PFKs grouped the enzymes into three monophyletic clusters: PFK group I represents only classical ATP-PFKs from Bacteria and Eucarya; PFK group II contains only PPi-PFKs from the genus Propionibacterium, plants, and amitochondriate protists; whereas group III consists of PFKs with either cosubstrate specificity, i.e., the PPi-dependent enzymes from T. tenax and Amycolatopsis methanolica and the ATP-PFK from Streptomyces coelicolor. Comparative analyses of the pattern of conserved active-site residues strongly suggest that the group III PFKs originally bound PPi as a cosubstrate.
authors.affiliation: FB 9 Mikrobiologie, Universität GH Essen, Germany. bettina.siebers@uni-essen.de
authors.forename: B
authors.initials: B
authors.lastname: Siebers
authors.forename: H P
authors.initials: HP
authors.lastname: Klenk
authors.forename: R
authors.initials: R
authors.lastname: Hensel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001105:Archaea / Q000145:classification / Q000201:enzymology*; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D019143:Evolution, Molecular; D005798:Genes, Bacterial; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010770:Phosphotransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D019624:Thermoproteaceae / Q000145:classification* / Q000201:enzymology* / Q000235:genetics
chemical_list: D046911:Macromolecular Substances; D011994:Recombinant Proteins; D010770:Phosphotransferases; C022502:pyrophosphate-fructose 6-phosphate 1-phosphotransferase