geranyl-pyrophosphate and isopentenyl-pyrophosphate

Farnesyl diphosphate synthase (FPPS) is an isoprenoid chain elongation enzyme that catalyzes the sequential condensation of dimethylallyl diphosphate (C

Topics: Binding Sites; Crystallography, X-Ray; Diphosphates; Diterpenes; Geranyltranstransferase; Hemiterpenes; Humans; Leishmania major; Leishmaniasis, Cutaneous; Models, Molecular; Organophosphorus Compounds; Polyisoprenyl Phosphates; Protein Conformation; Sesquiterpenes; Substrate Specificity

In this study, nerol was biosynthesized in the metabolic engineered Escherichia coli from glucose for the first time. Firstly, the truncated neryl diphosphate synthase gene tNDPS1 was expressed that catalyzes isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) to form neryl diphosphate (NPP), and then the nerol synthase gene GmNES was co-expressed to synthesize the final product nerol from NPP. The engineered strain LZ001 accumulated 0.053 ± 0.015 mg/L of nerol. Secondly, the IDI1, MVD1, ERG8, ERG12, tHMG1 and ERG13 were co-expressed to increase the supply of IPP and DMAPP. Finally, the heterologous ERG10 gene was overexpressed, and the recombinant strain LZ005 produced 1.564 ± 0.102 mg/L of nerol in shaking-flask culture, which represents a 29.51-fold increase over LZ001 strain. This study shows the novel method for the biosynthesis of nerol and provides new metabolic engineering strategy for the production of terpenoids.

Topics: Acyclic Monoterpenes; Escherichia coli; Glucose; Hemiterpenes; Metabolic Engineering; Organophosphorus Compounds; Polyisoprenyl Phosphates; Terpenes

Topics: Acyclic Monoterpenes; Camptotheca; Diphosphates; Diterpenes; DNA, Complementary; Escherichia coli; Geranyltranstransferase; Hemiterpenes; Monoterpenes; Organophosphorus Compounds; Polyisoprenyl Phosphates; Polymerase Chain Reaction; Sesquiterpenes; Terpenes

Farnesyl diphosphate synthase (FPPS) is a key enzyme responsible for the supply of isoprenoid precursors for several essential metabolites, including sterols, dolichols and ubiquinone. In Saccharomyces cerevisiae, FPPS catalyzes the sequential condensation of two molecules of isopentenyl diphosphate (IPP) with dimethylallyl diphosphate (DMAPP), producing geranyl diphosphate (GPP) and farnesyl diphosphate (FPP). Critical amino acid residues that determine product chain length were determined by a comparative study of strict GPP synthases versus strict FPPS. In silico ΔΔG, i.e. differential binding energy between a protein and two different ligands-of yeast FPPS mutants was evaluated, and F96, A99 and E165 residues were identified as key determinants for product selectivity. A99X variants were evaluated in vivo, S. cerevisiae strains carrying A99R and A99H variants showed significant differences on GPP concentrations and specific growth rates. The FPPS A99T variant produced unquantifiable amounts of FPP and no effect on GPP production was observed. Strains carrying A99Q, A99Y and A99K FPPS accumulated high amounts of DMAPP-IPP, with a decrease in GPP and FPP. Our results demonstrated the relevance of the first residue before FARM (First Aspartate Rich Motif) over substrate consumption and product specificity of S. cerevisiae FPPS in vivo. The presence of A99H significantly modified product selectivity and appeared to be relevant for GPP synthesis.

Topics: Amino Acid Motifs; Amino Acid Sequence; Amino Acid Substitution; Binding Sites; Diphosphates; Diterpenes; Gene Expression Regulation, Fungal; Geranyltranstransferase; Hemiterpenes; Kinetics; Metabolic Engineering; Molecular Docking Simulation; Organophosphorus Compounds; Point Mutation; Polyisoprenyl Phosphates; Protein Binding; Protein Interaction Domains and Motifs; Protein Structure, Secondary; Saccharomyces cerevisiae; Sequence Alignment; Sequence Homology, Amino Acid; Sesquiterpenes; Substrate Specificity; Terpenes; Thermodynamics

Subcellular monoterpene biosynthesis capacity based on local geranyl diphosphate (GDP) availability or locally boosted GDP production was determined for plastids, cytosol and mitochondria. A geraniol synthase (GES) was targeted to plastids, cytosol, or mitochondria. Transient expression in Nicotiana benthamiana indicated local GDP availability for each compartment but resulted in different product levels. A GDP synthase from Picea abies (PaGDPS1) was shown to boost GDP production. PaGDPS1 was also targeted to plastids, cytosol or mitochondria and PaGDPS1 and GES were coexpressed in all possible combinations. Geraniol and geraniol-derived products were analyzed by GC-MS and LC-MS, respectively. GES product levels were highest for plastid-targeted GES, followed by mitochondrial- and then cytosolic-targeted GES. For each compartment local boosting of GDP biosynthesis increased GES product levels. GDP exchange between compartments is not equal: while no GDP is exchanged from the cytosol to the plastids, 100% of GDP in mitochondria can be exchanged to plastids, while only 7% of GDP from plastids is available for mitochondria. This suggests a direct exchange mechanism for GDP between plastids and mitochondria. Cytosolic PaGDPS1 competes with plastidial GES activity, suggesting an effective drain of isopentenyl diphosphate from the plastids to the cytosol.

Topics: Acyclic Monoterpenes; Cytosol; Diphosphates; Diterpenes; Geranyltranstransferase; Hemiterpenes; Mitochondria; Monoterpenes; Nicotiana; Organophosphorus Compounds; Phosphoric Monoester Hydrolases; Picea; Plant Proteins; Plants, Genetically Modified; Plastids; Terpenes; Valerian

Terpenoids, compounds found in all domains of life, represent the largest class of natural products with essential roles in their hosts. All terpenoids originate from the five-carbon building blocks, isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP), which can be derived from the mevalonic acid (MVA) and methylerythritol phosphate (MEP) pathways. The absence of two components of the MVA pathway from archaeal genomes led to the discovery of an alternative MVA pathway with isopentenyl phosphate kinase (IPK) catalyzing the final step, the formation of IPP. Despite the fact that plants contain the complete classical MVA pathway, IPK homologs were identified in every sequenced green plant genome. Here, we show that IPK is indeed a member of the plant terpenoid metabolic network. It is localized in the cytosol and is coexpressed with MVA pathway and downstream terpenoid network genes. In planta, IPK acts in parallel with the MVA pathway and plays an important role in regulating the formation of both MVA and MEP pathway-derived terpenoid compounds by controlling the ratio of IP/DMAP to IPP/DMAPP. IP and DMAP can also competitively inhibit farnesyl diphosphate synthase. Moreover, we discovered a metabolically available carbon source for terpenoid formation in plants that is accessible via IPK overexpression. This metabolite reactivation approach offers new strategies for metabolic engineering of terpenoid production.

Topics: Arabidopsis; Arabidopsis Proteins; Archaea; Cytosol; Gene Expression Regulation, Plant; Gene Knockout Techniques; Genes, Plant; Hemiterpenes; Kinetics; Metabolic Networks and Pathways; Mevalonic Acid; Nicotiana; Organophosphorus Compounds; Phosphotransferases (Alcohol Group Acceptor); Plants, Genetically Modified; Plastids; Polyisoprenyl Phosphates; Sequence Homology, Amino Acid; Sesquiterpenes; Terpenes

A structure-based approach was pursued in designing novel bisphosphonate inhibitors of the human farnesyl pyrophosphate synthase (hFPPS). Preliminary SAR and structural evidence for the simultaneous binding of these inhibitors into the isopentenyl pyrophosphate (IPP) and the geranyl pyrophosphate (GPP) substrate sub-pockets of the enzyme are presented.

Topics: Binding Sites; Computer Simulation; Diphosphonates; Enzyme Inhibitors; Etidronic Acid; Geranylgeranyl-Diphosphate Geranylgeranyltransferase; Hemiterpenes; Humans; Organophosphorus Compounds; Polyisoprenyl Phosphates; Protein Binding; Risedronic Acid; Structure-Activity Relationship

Compounds of the mevalonate pathway containing a terminal di- or triphosphate (mev-PP or mev-PPP) were tested as substrates of several enzyme ligases (T4 RNA ligase, T4 DNA ligase, firefly luciferase and other ligases) for the synthesis of ATP derivatives of the mev-pppA or mev-ppppA type. T4 RNA ligase, in the presence of ATP and the substrates: geranyl, farnesyl or isopentenyl triphosphates, and geranyl, farnesyl, dimethylallyl or isopentenyl diphosphates, all at 0.3 mM concentration, catalyzed the synthesis of the corresponding ATP derivatives at a relative rate of activity of: 7.6+/-1.4 mU/mg or 100%; 39%; 42%; 24%; 18%; 12% and 6%, respectively. Inhibition (%) of the synthesis by excess of substrate (0.8 mM vs. 0.3 mM) was observed with farnesyl diphosphate (99%); farnesyl triphosphate (96%) and geranyl triphosphate (32%). V(max), K(m), K(cat) and K(cat)/K(m) values were also determined. The K(cat)/K(m) values calculated were for: farnesyl triphosphate, 166; geranyl triphosphate, 52.2; farnesyl diphosphate, 12.1; geranyl diphosphate, 8.6; isopentenyl triphosphate, 6.7; dimethylallyl diphosphate, 3.1 and isopentenyl diphosphate, 0.9. Similar results were obtained with T4 DNA ligase. The above-mentioned compounds were also substrates of firefly luciferase synthesizing the mev-pppA or mev-ppppA derivatives. In our hands, neither the acyl- or acetyl-CoA synthetases nor the ubiquiting activating enzyme (E1) catalyzed the synthesis of ATP derivatives of these compounds. The results here presented could be related with the mechanism of action of bisphosphonates on osteoclasts or tumor cells.

Topics: Adenosine Triphosphate; Animals; Binding Sites; Chromatography, High Pressure Liquid; Diphosphates; Diphosphonates; Diterpenes; DNA Ligases; Hemiterpenes; Mevalonic Acid; Organophosphorus Compounds; Osteoclasts; Polyisoprenyl Phosphates; Polyphosphates; RNA Ligase (ATP); Sesquiterpenes; Substrate Specificity

We identified a cis-prenyltransferase gene, neryl diphosphate synthase 1 (NDPS1), that is expressed in cultivated tomato (Solanum lycopersicum) cultivar M82 type VI glandular trichomes and encodes an enzyme that catalyzes the formation of neryl diphosphate from isopentenyl diphosphate and dimethylallyl diphosphate. mRNA for a terpene synthase gene, phellandrene synthase 1 (PHS1), was also identified in these glands. It encodes an enzyme that uses neryl diphosphate to produce beta-phellandrene as the major product as well as a variety of other monoterpenes. The profile of monoterpenes produced by PHS1 is identical with the monoterpenes found in type VI glands. PHS1 and NDPS1 map to chromosome 8, and the presence of a segment of chromosome 8 derived from Solanum pennellii LA0716 causes conversion from the M82 gland monoterpene pattern to that characteristic of LA0716 plants. The data indicate that, contrary to the textbook view of geranyl diphosphate as the "universal" substrate of monoterpene synthases, in tomato glands neryl diphosphate serves as a precursor for the synthesis of monoterpenes.

Topics: Alkyl and Aryl Transferases; Biosynthetic Pathways; Cloning, Molecular; Cyclohexane Monoterpenes; Dimethylallyltranstransferase; Diphosphates; Diterpenes; DNA, Complementary; Gas Chromatography-Mass Spectrometry; Gene Expression Regulation, Plant; Gene Library; Hemiterpenes; Kinetics; Molecular Sequence Data; Molecular Structure; Monoterpenes; Organophosphorus Compounds; Plant Epidermis; Plant Proteins; Polyisoprenyl Phosphates; Reverse Transcriptase Polymerase Chain Reaction; Sequence Analysis, DNA; Solanum lycopersicum; Substrate Specificity

Topics: Alkyl and Aryl Transferases; Dimethylallyltranstransferase; Hemiterpenes; Monoterpenes; Organophosphorus Compounds; Plant Epidermis; Plant Proteins; Polyisoprenyl Phosphates; Solanum lycopersicum; Substrate Specificity

Coenzyme Q(10) (CoQ(10)), like other CoQs of various organisms, plays indispensable roles not only in energy generation but also in several other processes required for cells' survival. In this study, a gene encoding for a decaprenyl diphosphate synthase (Rsdds) was cloned from Rhodobacter sphaeroides in Escherichia coli. The in vivo catalytic activity and product specificity of Rsdds were compared with those of a counterpart enzyme from Agrobacterium tumefaciens (Atdds) in E. coli as a heterologous host. In contrast with Atdds, Rsdds showed lower catalytic activity but higher product specificity for CoQ(10) production, as indicated by the amount of CoQ(9) formation. The higher product specificity of Rsdds was also confirmed by utilizing both Rsdds and Atdds for in vitro synthesis of polyprenyl diphosphates. Thin layer chromatography indicated that the Rsdds enzyme resulted in relatively much less solanesyl diphosphate formation. The purified Rsdds catalyzed the addition of isopentenyl diphosphate to dimethyl allyl diphosphate, geranyl diphosphate, omega,E,E-farnesyl diphosphate (FPP), and omega,E,E,E-geranylgeranyl diphosphate as priming substrates. The kinetic parameters of V (max) (pmol/min), K (M) (microM), k (cat) (1/min), and k (cat) /K (M) of the enzyme using FPP as the most appropriate substrate were determined to be 264.6, 13.1, 8.8, and 0.67, respectively.

Topics: Agrobacterium tumefaciens; Alkyl and Aryl Transferases; Chromatography, Thin Layer; Cloning, Molecular; Coenzymes; Diphosphates; Diterpenes; DNA, Bacterial; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Hemiterpenes; Kinetics; Molecular Sequence Data; Organophosphorus Compounds; Polyisoprenyl Phosphates; Rhodobacter sphaeroides; Sequence Analysis, DNA; Sesquiterpenes; Substrate Specificity; Ubiquinone

UPPS (undecaprenyl pyrophosphate synthase) catalyses consecutive condensation reactions of FPP (farnesyl pyrophosphate) with eight isopentenyl pyrophosphates to generate C55 UPP, which serves as a lipid carrier for bacterial peptidoglycan biosynthesis. We reported the co-crystal structure of Escherichia coli UPPS in complex with FPP. Its phosphate head-group is bound to positively charged arginine residues and the hydrocarbon moiety interacts with hydrophobic amino acids including L85, L88 and F89, located on the alpha3 helix of UPPS. We now show that the monophosphate analogue of FPP binds UPPS with an eight times lower affinity (K(d)=4.4 microM) compared with the pyrophosphate analogue, a result of a larger dissociation rate constant (k(off)=192 s(-1)). Farnesol (1 mM) lacking the pyrophosphate does not inhibit the UPPS reaction. GGPP (geranylgeranyl pyrophosphate) containing a larger C20 hydrocarbon tail is an equally good substrate (K(m)=0.3 microM and kcat=2.1 s(-1)) compared with FPP. The shorter C10 GPP (geranyl pyrophosphate) displays a 90-fold larger K(m) value (36.0+/-0.1 microM) but similar kcat value (1.7+/-0.1 s(-1)) compared with FPP. Replacement of L85, L88 or F89 with Ala increases FPP and GGPP K(m) values by the same amount, indicating that these amino acids are important for substrate binding, but do not determine substrate specificity. With GGPP as a substrate, UPPS still catalyses eight isopentenyl pyrophosphate condensation reactions to synthesize C60 product. Computer modelling suggests that the upper portion of the active-site tunnel, where cis double bonds of the product reside, may be critical for determining the final product chain length.

Topics: Alkyl and Aryl Transferases; Binding Sites; Escherichia coli Proteins; Farnesol; Hemiterpenes; Hydrophobic and Hydrophilic Interactions; Kinetics; Models, Molecular; Molecular Weight; Mutagenesis, Site-Directed; Organophosphorus Compounds; Polyisoprenyl Phosphates; Protein Conformation; Recombinant Fusion Proteins; Sesquiterpenes; Substrate Specificity

Cell-free homogenates from sage (Salvia officinalis) leaves convert dimethylallyl pyrophosphate and isopentenyl pyrophosphate to a mixture of geranyl pyrophosphate, farnesyl pyrophosphate, and geranylgeranyl pyrophosphate, with farnesyl pyrophosphate predominating. These prenyltransferase activities were localized primarily in the soluble enzyme fraction, and separation of this preparation on Sephadex G-150 revealed the presence of a partially resolved, labile geranyl pyrophosphate synthase activity. The product of the condensation reaction between [1-14C]dimethylallyl pyrophosphate and [1-3H]isopentenyl pyrophosphate was verified as [14C,1-3H]geranyl pyrophosphate by TLC isolation, enzymatic hydrolysis to geraniol, degradative studies, and the preparation of the crystalline diphenylurethane. The cis-isomer, neryl pyrophosphate, was not a product of the enzymatic reaction. By employing a selective tissue extraction procedure, the geranyl pyrophosphate synthase activity was localized in the leaf epidermal glands, the site of monoterpene biosynthesis, suggesting that the role of this enzyme is to supply the C10 precursor for the production of monoterpenes. Glandular extracts enriched in geranyl pyrophosphate synthase were partially purified by a combination of hydrophobic interaction chromatography on phenyl-Sepharose and gel permeation chromatography on Sephadex G-150. Substrate and product specificity studies confirmed the selective synthesis of geranyl pyrophosphate by this enzyme, which was also characterized with respect to molecular weight, pH optimum, cation requirement, inhibitors, and kinetic parameters, and shown to resemble other prenyltransferases.

Topics: Cell-Free System; Chromatography; Chromatography, Thin Layer; Dimethylallyltranstransferase; Hemiterpenes; Hydrogen-Ion Concentration; Organophosphorus Compounds; Plants; Polyisoprenyl Phosphates; Sesquiterpenes; Terpenes; Transferases

A prenyltransferase purified from the commercial rubber tree, Hevea brasiliensis, that elongates existing cis-polyisoprene rubber molecules also catalyzes the formation of all trans-farnesyl pyrophosphate (t,t-FPP) from dimethylallyl pyrophosphate (DMAPP) and isopentenyl pyrophosphate (IPP). In assays of the latter activity trans-geranyl pyrophosphate is the only other product identified. In contrast to this limited addition of IPP to DMAPP, we measured 7000 additions of isoprene per rubber molecule in a previous titration of active allylic ends of rubber molecules by purified prenyltransferase (Light, D. R., and Dennis, M. S. (1989) J. Biol. Chem. 264, 18589-18597). In order to confirm that purified prenyltransferase extensively elongates rubber molecules, doubly labeled [1-14C]isopentenyl [U-32P]pyrophosphate ([14C,32P]IPP) was synthesized. Using this reagent we show that both prenyltransferase purified from H. brasiliensis and prenyltransferase purified from avian liver (FPP synthase) add greater than 15 isoprene units to existing rubber molecules, consistent with the previous titration data. For confirmation that the prenyltransferase purified from H. brasiliensis adds isoprene units to rubber to make cis-polyisoprene, chirally tritiated [14C]IPP ([14C,2S-3H]IPP) was synthesized. Retention of the tritium label in FPP synthesized from [14C,2S-3H]IPP and DMAPP, geranyl pyrophosphate, or neryl pyrophosphate by prenyltransferase from H. brasiliensis or avian liver confirms trans addition to these substrates. In contrast, when [14C,2S-3H]IPP is incubated with serum-free rubber particles and prenyltransferase purified from H. brasiliensis, avian liver, or yeast, no tritium is incorporated into the rubber particles indicating cis addition. Thus, rubber particles have the ability to alter the stereoselective removal of the 2R-prochiral proton in favor of the removal of the 2S-prochiral proton. This apparent inversion of carbon 2 of IPP during the proton abstraction step by rubber particles represents a novel example of a switch in enzyme stereospecificity. In addition to being enzymatically similar to other prenyltransferases, rubber transferase also appears to be related immunologically to FPP synthases, since polyclonal antibodies to the H. brasiliensis prenyltransferase cross-react with the purified yeast prenyltransferase. In order to investigate potential primers of greater molecular weight than that of FPP, cis-undecaprenyl pyrophosphate (C55PP) was syn

Topics: Animals; Chickens; Dimethylallyltranstransferase; Hemiterpenes; Liver; Molecular Weight; Organophosphorus Compounds; Phosphates; Plants; Polyisoprenyl Phosphates; Rubber; Sesquiterpenes; Stereoisomerism; Substrate Specificity; Transferases; Trees

We have purified "rubber transferase" from latex of the commercial rubber tree Hevea brasiliensis and find that it is a dimer with a monomeric molecular mass of 38,000 Da, requires Mg2+, and is stabilized by thiols in agreement with studies of a partially purified preparation previously described (Archer, B. L., and Cockbain, E. G. (1969) Methods Enzymol. 15, 476-480). Greater than 90% of the [1-14C]isopentenyl pyrophosphate which is incorporated into deproteinated rubber particles by the purified prenyltransferase is added to high molecular mass polyisoprene (greater than 20,000 Da). Purified prenyltransferase and deproteinated rubber particles reconstitute 40-60% of the biosynthetic activity of whole latex in samples matched for rubber content. Incorporation is linear with added rubber particles up to at least 10 mg/ml rubber or 20 microM rubber molecules (based on a number average molecular mass of 500,000 Da). Prenyltransferase concentrations estimated in whole latex (0.37% or 160 nM) are sufficient to saturate all elongation sites in whole latex, and addition of purified prenyltransferase does not increase [1-14C]isopentenyl pyrophosphate incorporation. Deproteinated rubber particles can be titrated with the pure enzyme (Kd = 9 nM) demonstrating that the fraction of rubber molecules available for addition is low (approximately 0.01%). An estimated 7,000 isoprene units are added per complex at a rate of 1/s in a typical assay. Hevea prenyltransferase catalyzes the formation of cis-isoprene in the presence of rubber particles. However, in the absence of rubber particles and in the presence of dimethylallyl pyrophosphate, the purified prenyltransferase catalyzes the formation of geranyl pyrophosphate and all trans-farnesyl pyrophosphate as demonstrated by thin layer chromatography, gas chromatography, and molecular exclusion chromatography.

Topics: Amino Acid Sequence; Chromatography; Chromatography, High Pressure Liquid; Dimethylallyltranstransferase; Dithiothreitol; Hemiterpenes; Kinetics; Latex; Macromolecular Substances; Molecular Sequence Data; Molecular Weight; Organophosphorus Compounds; Plants; Polyisoprenyl Phosphates; Rubber; Sesquiterpenes; Substrate Specificity; Transferases; Trees

Upon rehydration of lyophilized Escherichia coli cells with phosphate buffer containing [14C]isopentenyl pyrophosphate (IPP), 14C was incorporated into the cells. Radioactivity was found in ubiquinone-8, an unidentified precursor of ubiquinone-8, demethylmenaquinone-8 and phosphate esters of all-trans-octaprenol and cis, trans-polyprenols. On rehydration of the cells with the buffer containing geranyl pyrophosphate or farnesyl pyrophosphate in combination with [14C]IPP, higher radioactivity was incorporated into the above products and some radioactivity was found in free prenols. Fractionation of the 14C-labeled cells by sucrose-density gradient centrifugation before and after recultivation indicated that the size of 14C-labeled cells had changed during the recultivation. This shows that radioactivity of [14C]IPP was incorporated into live cells but not into dead cells. The metabolism of the radioactive products in the recultivated cells was examined. It was found that the unidentified precursor was converted to ubiquinone-8, but demethylmenaquinone-8 was not converted to menaquinone-8. "Lipid intermediates" in peptidoglycan synthesis increased in the logarithmic growth phase and decreased in the stationary phase. In the stationary phase, however, an increase in cis,trans-polyprenyl monophosphates was observed. These observations suggest the operation of the lipid cycle of peptidoglycan synthesis.

Topics: Carbon Radioisotopes; Cells, Cultured; Centrifugation, Density Gradient; Chromatography, Thin Layer; Escherichia coli; Freeze Drying; Hemiterpenes; Isomerism; Organophosphorus Compounds; Polyisoprenyl Phosphates; Sesquiterpenes

Topics: Diphosphates; Hemiterpenes; Organophosphorus Compounds; Polyisoprenyl Phosphates; Sesquiterpenes

Farnesyl pyrophosphate synthetase was detected in extracts of Bacillus subtilis and partially purified by Sephadex G-100, hydroxylapatite, and DEAE-Sephadex chromatography. The enzyme catalyzed the exclusive formation of all-trans farnesyl pyrophosphate from isopentenyl pyrophosphate and either dimethylallyl or geranyl pyrophosphate. Mg2+ was essential for the catalytic activity and Mn2+ was less effective. The enzyme was slightly activated by sulfhydryl reagents. This enzyme was markedly stimulated by K+, NH4+, or detergents such as Triton X-100 and Tween 80, unlike the known farnesyl pyrophosphate synthetases from eucaryotes. The molecular weight of the enzyme was estimated by gel filtration to be 67,000. The Michaelis constants for dimethylallyl and geranyl pyrophosphate were 50 microM and 18 microM, respectively.

Topics: Alkenes; Bacillus subtilis; Bacitracin; Detergents; Dimethylallyltranstransferase; Farnesol; Hemiterpenes; Magnesium; Organophosphorus Compounds; Polyisoprenyl Phosphates; Sesquiterpenes; Sulfhydryl Reagents; Transferases