Home About us Contact
|
Home About us Contact | ||
|
|
||
Farnesyl Diphosphate (farnesyl + diphosphate)
Terms modified by Farnesyl Diphosphate Selected AbstractsProtein farnesyltransferase inhibitors interfere with farnesyl diphosphate binding by rubber transferaseFEBS JOURNAL, Issue 19 2003Christopher J. D. Mau Rubber transferase, a cis -prenyltransferase, catalyzes the addition of thousands of isopentenyl diphosphate (IPP) molecules to an allylic diphosphate initiator, such as farnesyl diphosphate (FPP, 1), in the presence of a divalent metal cofactor. In an effort to characterize the catalytic site of rubber transferase, the effects of two types of protein farnesyltransferase inhibitors, several chaetomellic acid A analogs (2, 4,7) and ,-hydroxyfarnesylphosphonic acid (3), on the ability of rubber transferase to add IPP to the allylic diphosphate initiator were determined. Both types of compounds inhibited the activity of rubber transferases from Hevea brasiliensis and Parthenium argentatum, but there were species,specific differences in the inhibition of rubber transferases by these compounds. Several shorter analogs of chaetomellic acid A did not inhibit rubber transferase activity, even though the analogs contained chemical features that are present in an elongating rubber molecule. These results indicate that the initiator-binding site in rubber transferase shares similar features to FPP binding sites in other enzymes. [source] Fusion of farnesyldiphosphate synthase and epi -aristolochene synthase, a sesquiterpene cyclase involved in capsidiol biosynthesis in Nicotiana tabacumFEBS JOURNAL, Issue 14 2002Maria Brodelius A clone encoding farnesyl diphosphate synthase (FPPS) was obtained by PCR from a cDNA library made from young leaves of Artemisia annua. A cDNA clone encoding the tobacco epi -aristolochene synthase (eAS) was kindly supplied by J. Chappell (University of Kentucky, Lexington, KY, USA). Two fusions were constructed, i.e. FPPS/eAS and eAS/FPPS. The stop codon of the N-terminal enzyme was removed and replaced by a short peptide (Gly-Ser-Gly) to introduce a linker between the two ORFs. These two fusions and the two single cDNA clones were separately introduced into a bacterial expression vector (pET32). Escherichia coli was transformed with the expression vectors and enzymatically active soluble proteins were obtained after induction with isopropyl thio-,- d -thiogalactoside. The recombinant enzymes were purified using immobilized metal affinity chromatography on Co2+ columns. The fusion enzymes produced epi- aristolochene from isopentenyl diphosphate through a coupled reaction. The Km values of FPPS and eAS for isopentenyl diphosphate and farnesyl diphosphate, respectively, were essentially the same for the single and fused enzymes. The bifunctional enzymes showed a more efficient conversion of isopentenyl diphosphate to epi -aristolochene than the corresponding amount of single enzymes. [source] The Intracellular Target for the Antiresorptive Aminobisphosphonate Drugs in Dictyostelium discoideum Is the Enzyme Farnesyl Diphosphate Synthase,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 5 2000Joanna E. Grove Abstract Aminobisphosphonate (aBP) drugs inhibit osteoclast-mediated bone resorption and also growth of amoebas of Dictyostelium discoideum apparently by interaction with the same intracellular target. Identification of the target in Dictyostelium therefore could also identify the target in osteoclasts. The aBPs (100 ,M alendronate and 30 ,M YM-175) inhibited conversion of [14C]mevalonate into sterols by cultures of Dictyostelium amoebas. One of three enzymes (isopentenyl diphosphate [IDP] isomerase, farnesyl diphosphate [FDP] synthase, and squalene synthase) appeared to be the target for this inhibition because conversion of [14C]IDP into squalene, the immediate precursor for sterol biosynthesis, was inhibited in extracts of wild-type amoebas by alendronate (IC50 = 75 nM) or risedronate (IC50 = 30 nM) whereas, when the extract had been prepared from amoebas of strains selected for having partial resistance to the growth-inhibitory effects of alendronate (strain MR102) or risedronate (strain RB101), the values of IC50 were increased to 700 nM for alendronate (MR102 extract) or 130 nM for risedronate (RB101 extract). Neither IDP isomerase nor squalene synthase was inhibited significantly by alendronate or risedronate but both of these aBP drugs, and all others tested, inhibited FDP synthase. Determination of the nucleotide sequences of complementary DNAs (cDNAs) encoding FDP synthase in the wild-type and aBP-resistant strains of Dictyostelium indicated that there had been no changes in the amino acid sequence of the enzyme in the mutant strains. However, both mutant strains overproduce FDP synthase. It is concluded that FDP synthase is the intracellular target for the aBP drugs. (J Bone Miner Res 2000;15:971,981) [source] A New Farnesyl Diphosphate Synthase Gene from Taxus media Rehder: Cloning, Characterization and Functional ComplementationJOURNAL OF INTEGRATIVE PLANT BIOLOGY, Issue 6 2006Zhi-Hua Liao Abstract Farnesyl diphosphate synthase (FPS; EC 2.5.1.10) catalyzes the production of 15-carbon farnesyl diphosphate which is a branch-point intermediate for many terpenoids. This reaction is considered to be a rate-limiting step in terpenoid biosynthesis. Here we report for the first time the cloning of a new full-length cDNA encoding farnesyl diphosphate synthase from a gymnosperm plant species, Taxus media Rehder, designated as TmFPS1. The full-length cDNA of TmFPS1 (GenBank accession number: AY461811) was 1 464 bp with a 1 056-bp open reading frame encoding a 351-amino acid polypeptide with a calculated molecular weight of 40.3 kDa and a theoretical pI of 5.07. Bioinformatic analysis revealed that TmFPS1 contained all five conserved domains of prenyltransferases, and showed homology to other FPSs of plant origin. Phylogenetic analysis showed that farnesyl diphosphate synthases can be divided into two groups: one of prokaryotic origin and the other of eukaryotic origin. TmFPS1 was grouped with FPSs of plant origin. Homology-based structural modeling showed that TmFPS1 had the typical spatial structure of FPS, whose most prominent structural feature is the arrangement of 13 core helices around a large central cavity in which the catalytic reaction takes place. Our bioinformatic analysis strongly suggests that TmFPS1 is a functional gene. Southern blot analysis revealed that TmFPS1 belongs to a small FPS gene family in T. media. Northern blot analysis indicated that TmFPS1 is expressed in all tested tissues, including the needles, stems and roots of T. media. Subsequently, functional complementation with TmFPS1 in a FPS-deficient mutant yeast demonstrated that TmFPS1 did encode farnesyl diphosphate synthase, which rescued the yeast mutant. This study will be helpful in future investigations aiming at understanding the detailed role of FPS in terpenoid biosynthesis flux control at the molecular genetic level. (Managing editor: Wei Wang) [source] Isolation and characterization of farnesyl diphosphate synthase from the cotton boll weevil, Anthonomus grandisARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 2 2009A. Huma Taban Abstract Farnesyl diphosphate synthase (FPPS) catalyzes the consecutive condensation of two molecules of isopentenyl diphosphate with dimethylallyl diphosphate to form farnesyl diphosphate (FPP). In insects, FPP is used for the synthesis of ubiquinones, dolicols, protein prenyl groups, and juvenile hormone. A full-length cDNA of FPPS was cloned from the cotton boll weevil, Anthonomus grandis (AgFPPS). AgFPPS cDNA consists of 1,835 nucleotides and encodes a protein of 438 amino acids. The deduced amino acid sequence has high similarity to previously isolated insect FPPSs and other known FPPSs. Recombinant AgFPPS expressed in E. coli converted labeled isopentenyl diphosphate in the presence of dimethylallyl diphosphate to FPP. Southern blot analysis indicated the presence of a single copy gene. Using molecular modeling, the three-dimensional structure of coleopteran FPPS was determined and compared to the X-ray crystal structure of avian FPPS. The ,-helical fold is conserved in AgFPPS and the size of the active site cavity is consistent with the enzyme being a FPPS. © 2009 Wiley Periodicals, Inc. [source] Enhancement of farnesyl diphosphate pool as direct precursor of sesquiterpenes through metabolic engineering of the mevalonate pathway in Saccharomyces cerevisiaeBIOTECHNOLOGY & BIOENGINEERING, Issue 1 2010Mohammad A. Asadollahi Abstract The mevalonate pathway in the yeast Saccharomyces cerevisiae was deregulated in order to enhance the intracellular pool of farnesyl diphosphate (FPP), the direct precursor for the biosynthesis of sesquiterpenes. Over-expression of the catalytic domain of HMG1, both from the genome and plasmid, resulted in higher production of cubebol, a plant originating sesquiterpene, and increased squalene accumulation. Down-regulation of ERG9 by replacing its native promoter with the regulatable MET3 promoter, enhanced cubebol titers but simultaneous over-expression of tHMG1 and repression of ERG9 did not further improve cubebol production. Furtheremore, the concentrations of squalene and ergosterol were measured in the engineered strains. Unexpectedly, significant accumulation of squalene and restoring the ergosterol biosynthesis were observed in the ERG9 repressed strains transformed with the plasmids harboring cubebol synthase gene. This could be explained by a toxicity effect of cubebol, possibly resulting in higher transcription levels for the genes under control of MET3 promoter, which could lead to accumulation of squalene and ergosterol. Biotechnol. Bioeng. 2010; 106: 86,96. © 2010 Wiley Periodicals, Inc. [source] The in vivo synthesis of plant sesquiterpenes by Escherichia coliBIOTECHNOLOGY & BIOENGINEERING, Issue 5 2001Vincent J.J. Martin Abstract Three plant genes encoding (+)-,-cadinene, 5- epi -aristolochene, and vetispiradiene cyclases were expressed in Escherichia coli to evaluate the potential of this bacterium to synthesize sesquiterpenes in vivo. Various growth temperatures, carbon sources, and host strains were examined to optimize terpene production. The highest levels of sesquiterpene production occurred when the enzymes were expressed in strain DH5, from the trc promoter (Ptrc) of the high-copy plasmidpTrc99A in M9 medium supplemented with 0.2% (v/v) glycerol at 30°C for 5- epi -aristolochene and vetispiradiene and 37°C for (+)-,-cadinene. The highest concentrations of sesquiterpenes observed were 10.3 ,g of (+)-,-cadinene, 0.24 ,g of 5- epi -aristolochene (measured as (+)-,-cadinene equivalents), and 6.4 ,g of vetispiradiene (measured as (+)-,-cadinene equivalents) per liter of culture. These sesquiterpene production levels are >500-fold lower than carotenoid production, both of which are synthesized from endogenous trans -farnesyl diphosphate (FDP) in E. coli. Based on these results, we conclude that the limiting factor for sesquiterpene synthesis in E. coli is the poor expression of the cyclase enzyme and not supply of the FDP precursor. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 75: 497,503, 2001. [source] | |||||||||||||