title: The farnesyl-diphosphate/geranylgeranyl-diphosphate synthase of Toxoplasma gondii is a bifunctional enzyme and a molecular target of bisphosphonates.
abstract: Farnesyl-diphosphate synthase (FPPS) catalyzes the synthesis of farnesyl diphosphate, an important precursor of sterols, dolichols, ubiquinones, and prenylated proteins. We report the cloning and characterization of two Toxoplasma gondii farnesyl-diphosphate synthase (TgFPPS) homologs. A single genetic locus produces two transcripts, TgFPPS and TgFPPSi, by alternative splicing. Both isoforms were heterologously expressed in Escherichia coli, but only TgFPPS was active. The protein products predicted from the nucleotide sequences have 646 and 605 amino acids and apparent molecular masses of 69.5 and 64.5 kDa, respectively. Several conserved sequence motifs found in other prenyl-diphosphate synthases are present in both TgFPPSs. TgFPPS was also expressed in the baculovirus system and was biochemically characterized. In contrast to the FPPS of other eukaryotic organisms, TgFPPS is bifunctional, catalyzing the formation of both farnesyl diphosphate and geranylgeranyl diphosphate. TgFPPS localizes to the mitochondria, as determined by the co-localisation of the affinity-purified antibodies against the protein with MitoTracker, and in accord with the presence of an N-terminal mitochondria-targeting signal in the protein. This enzyme is an attractive target for drug development, because the order of inhibition of the enzyme by a number of bisphosphonates is the same as that for inhibition of parasite growth. In summary, we report the first bifunctional farnesyl-diphosphate/geranylgeranyl-diphosphate synthase identified in eukaryotes, which, together with previous results, establishes this enzyme as a valid target for the chemotherapy of toxoplasmosis.
authors.affiliation: Department of Pathobiology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Ling
authors.forename: Zhu-Hong
authors.initials: ZH
authors.lastname: Li
authors.forename: Kildare
authors.initials: K
authors.lastname: Miranda
authors.forename: Eric
authors.initials: E
authors.lastname: Oldfield
authors.forename: Silvia N J
authors.initials: SN
authors.lastname: Moreno
mesh_terms: D017398:Alternative Splicing / Q000187:drug effects / Q000502:physiology; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae; D050071:Bone Density Conservation Agents / Q000494:pharmacology / Q000627:therapeutic use; D002384:Catalysis; D003001:Cloning, Molecular; D011756:Diphosphates / Q000378:metabolism; D004164:Diphosphonates / Q000494:pharmacology* / Q000627:therapeutic use; D004224:Diterpenes / Q000378:metabolism; D004286:Dolichols / Q000096:biosynthesis; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000494:pharmacology* / Q000627:therapeutic use; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D051231:Farnesyltranstransferase / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D051228:Geranyltranstransferase / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011106:Polyisoprenyl Phosphates / Q000096:biosynthesis; D017368:Protein Prenylation / Q000187:drug effects / Q000502:physiology; D015800:Protozoan Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics* / Q000378:metabolism; D040641:Quantitative Trait Loci / Q000502:physiology; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012717:Sesquiterpenes; D013261:Sterols / Q000096:biosynthesis; D014122:Toxoplasma / Q000201:enzymology* / Q000235:genetics; D014123:Toxoplasmosis / Q000178:diet therapy / Q000201:enzymology / Q000235:genetics; D014451:Ubiquinone / Q000031:analogs & derivatives / Q000096:biosynthesis
chemical_list: D050071:Bone Density Conservation Agents; D011756:Diphosphates; D004164:Diphosphonates; D004224:Diterpenes; D004286:Dolichols; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D011106:Polyisoprenyl Phosphates; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D012717:Sesquiterpenes; D013261:Sterols; C511282:geranyl diphosphate; D014451:Ubiquinone; C004808:farnesyl pyrophosphate; D051228:Geranyltranstransferase; D051231:Farnesyltranstransferase; C003741:ubiquinol