Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The purification of 3,3-dimethylallyl- and geranyl-transferase and of isopentenyl pyrophosphate isomerase from pig liver.

ABSTRACT: The enzyme catalysing the synthesis of farnesyl pyrophosphate from dimethylallyl pyrophosphate and isopentenyl pyrophosphate, or from geranyl pyrophosphate and isopentenyl pyrophosphate, has been purified 100-fold from homogenates of pig liver. The enzyme has optimum pH 7.9 and requires Mg(2+) as activator in preference to Mn(2+); it is inhibited by iodoacetamide, N-ethylmaleimide, p-hydroxymercuribenzoate and phosphate ions in addition to the products of the reaction, inorganic pyrophosphate and farnesyl pyrophosphate. From product-inhibition studies of the geranyltransferase reaction, the order of addition of substrates to and release of products from the enzyme has been deduced: geranyl pyrophosphate combines with the enzyme first, followed by isopentenyl pyrophosphate. Farnesyl pyrophosphate dissociates from the enzyme before inorganic pyrophosphate. The existence of isopentenyl pyrophosphate isomerase in liver is confirmed. Methods for the preparation of the pyrophosphate esters of isopentenol, 3,3-dimethylallyl alcohol, geraniol and farnesol are also described.

SUBMITTER: Holloway PW

PROVIDER: S-EPMC1270545 | biostudies-other | 1967 Jul

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Isopentenyl pyrophosphate isomerase from liver.

Project description:Isopentenyl pyrophosphate isomerase (EC 5.3.3.2) was purified from extracts of pig liver by ammonium sulphate fractionation and by gel filtration. After about 20-fold purification the preparations were free of phosphatase and prenyltransferase (EC 2.5.1.1), the two enzymes that could have interfered with the assays. The isomerase has a distinct pH optimum at 6.0 and is activated by Mn(2+) in preference to Mg(2+). The K(m) value for isopentenyl pyrophosphate is 4x10(-6)m. The equilibrium of the reaction favours the formation of dimethylallyl pyrophosphate. The reversibility of the isomerase reaction was demonstrated directly by the formation of isopentenyl pyrophosphate from dimethylallyl pyrophosphate. It is suggested that two prenyl isomerases might exist, one involved in the synthesis of trans- and another in the synthesis of cis-polyprenyl substances.

| S-EPMC1198587 | biostudies-other

Identification and functional analysis of isopentenyl pyrophosphate isomerase genes in the whiteflies Bemisia tabaci (Hemiptera: Aleyrodidae).

Project description:The juvenile hormone (JH) plays a vital role in the regulation of a number of physiological processes, including development, reproduction, and ovarian maturation. Isopentenyl pyrophosphate isomerase (IPPI) is a key enzyme in the biosynthetic pathway of JH. In this study, we identified an isopentenyl pyrophosphate isomerase protein from Bemisia tabaci and named it BtabIPPI. The open reading frame (ORF) of BtabIPPI is 768 bp and encodes a protein of 255 amino acids that contains a conserved domain of the Nudix family. The temporal and spatial expression profiles showed that BtabIPPI was highly expressed in the female adults.RNA interference (RNAi)-mediated silencing of BtabIPPI reduced JH titers and the relative expression of vitellogenin receptor (VgR) and JH signaling pathway genes, resulting in a dramatic reduction in fecundity and hatchability. These results indicate that the BtabIPPI gene plays an important role in the female fecundity of B. tabaci. This study will broaden our understanding of the function of IPPI in regulating insect reproduction and provide a theoretical basis for targeting IPPI for pest control in the future.

| S-EPMC10278561 | biostudies-literature

Crystal structure of type 2 isopentenyl diphosphate isomerase from Thermus thermophilus in complex with inorganic pyrophosphate.

Project description:The N-terminal region is stabilized in the crystal structure of Thermus thermophilus type 2 isopentenyl diphosphate isomerase in complex with inorganic pyrophosphate, providing new insights about the active site and the catalytic mechanism of the enzyme. The PP i moiety is located near the conserved residues, H10, R97, H152, Q157, E158, and W219, and the flavin cofactor. The putative active site of isopentenyl diphosphate isomerase 2 provides interactions for stabilizing a carbocationic intermediate similar to those that stabilize the intermediate in the well-established protonation-deprotonation mechanism of isopentenyl diphosphate isomerase 1.

| S-EPMC4459642 | biostudies-literature

An improved synthesis of isopentenyl pyrophosphate.

Project description:1. Isopentenol was converted into isopentenyl phosphate with phosphoryl chloride in ether containing pyridine. 2. The isopentenyl phosphate reacted in 2-methylpropan-2-ol-water with morpholine and dicyclohexylcarbodi-imide to give isopentenyl phosphoromorpholidate. 3. The isopentenyl phosphoromorpholidate, with inorganic phosphate in pyridine containing tributylamine, gave isopentenyl pyrophosphate. The yield of pyrophosphate from monophosphate was 80-85% and the yield of pyrophosphate from isopentenol 40-60%. [1-(14)C]-Isopentenyl pyrophosphate was prepared by this method. 4. The yield of isopentenyl pyrophosphate from isopentenyl phosphate was substantially improved, in comparison with the yields obtained by published methods via the phosphoramidate, by the use of the phosphoromorpholidate.

| S-EPMC1198343 | biostudies-other

Crystal structure of isopentenyl diphosphate:dimethylallyl diphosphate isomerase.

Project description:Isopentenyl diphosphate:dimethylallyl diphosphate (IPP:DMAPP) isomerase catalyses a crucial activation step in the isoprenoid biosynthesis pathway. This enzyme is responsible for the isomerization of the carbon-carbon double bond of IPP to create the potent electrophile DMAPP. DMAPP then alkylates other molecules, including IPP, to initiate the extraordinary variety of isoprenoid compounds found in nature. The crystal structures of free and metal-bound Escherichia coli IPP isomerase reveal critical active site features underlying its catalytic mechanism. The enzyme requires one Mn(2+) or Mg(2+) ion to fold in its active conformation, forming a distorted octahedral metal coordination site composed of three histidines and two glutamates and located in the active site. Two critical residues, C67 and E116, face each other within the active site, close to the metal-binding site. The structures are compatible with a mechanism in which the cysteine initiates the reaction by protonating the carbon-carbon double bond, with the antarafacial rearrangement ultimately achieved by one of the glutamates involved in the metal coordination sphere. W161 may stabilize the highly reactive carbocation generated during the reaction through quadrupole- charge interaction.

| S-EPMC145486 | biostudies-literature

Isopentenyl diphosphate isomerase. Mechanism-based inhibition by diene analogues of isopentenyl diphosphate and dimethylallyl diphosphate.

Project description:Isopentenyl diphosphate isomerase (IDI) catalyzes the interconversion of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). This is an essential step in the mevalonate entry into the isoprenoid biosynthetic pathway. The isomerization catalyzed by type I IDI involves protonation of the carbon-carbon double bond in IPP or DMAPP to form a tertiary carbocation, followed by deprotonation. Diene analogues for DMAPP (E-2-OPP and Z-2-OPP) and IPP (4-OPP) were synthesized and found to be potent active-site-directed irreversible inhibitors of the enzyme. X-ray analysis of the E.I complex between Escherichia coli IDI and 4-OPP reveals the presence of two isomers that differ in the stereochemistry of the newly formed C3-C4 double bond in the hydrocarbon chain of the inhibitor. In both adducts C5 of the inhibitor is joined to the sulfur of C67. In these structures the methyl group formed upon protonation of the diene moiety in 4-OPP is located near E116, implicating that residue in the protonation step.

| S-EPMC2528281 | biostudies-literature

Integrated analysis of isopentenyl pyrophosphate (IPP) toxicity in isoprenoid-producing Escherichia coli

Project description:Isopentenyl pyrophosphate (IPP) is the universal C5 precursor for isoprenoids, the largest class of natural products and frequent targets for metabolic engineering. In engineered microbial systems, IPP toxicity presents a challenge since its formation is unavoidable in the production of high-value, long-chain terpenes. In this work, we develop an experimental platform to study IPP toxicity in E. coli engineered to produce isoprenol, an IPP-derived C5 alcohol. We first characterize the physiological response to IPP accumulation, demonstrating that elevated levels of IPP are linked to growth inhibition, altered morphology, and reduced cell viability. We show that IPP toxicity selects for pathway “breakage”, using proteomics to identify a reduction in phosphomevalonate kinase (PMK) as a probable recovery mechanism. Next, we demonstrate that endogenous E. coli metabolism is globally impacted by IPP accumulation, which results in inhibited nutrient uptake, reduced ATP levels, and an apparent “pause” in metabolism. Finally, we suggest that IPP toxicity is mediated by the formation of an isoprenyl-ATP analog (ApppI). The complementary data presented here represent the most comprehensive assessment of IPP stress to date and suggest potential strategies for the alleviation of IPP and prenyl diphosphate toxicity.

2018-03-14 | GSE102672 | GEO

Type-2 isopentenyl diphosphate isomerase: evidence for a stepwise mechanism.

Project description:Isopentenyl diphosphate isomerase (IDI) catalyzes the interconversion of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). These two molecules are the building blocks for construction of isoprenoid carbon skeletons in nature. Two structurally unrelated forms of IDI are known. A variety of studies support a proton addition/proton elimination mechanism for both enzymes. During studies with Thermus thermophilus IDI-2, we discovered that the olefinic hydrogens of a vinyl thiomethyl analogue of isopentenyl diphosphate exchanged with solvent when the enzyme was incubated with D(2)O without concomitant isomerization of the double bond. These results suggest that the enzyme-catalyzed isomerization reaction is not concerted.

| S-EPMC3237117 | biostudies-literature

Type-2 isopentenyl diphosphate isomerase. Mechanistic studies with cyclopropyl and epoxy analogues.

Project description: Not available

| S-EPMC2494526 | biostudies-literature

Reconstitution and substrate specificity for isopentenyl pyrophosphate of the antiviral radical SAM enzyme viperin.

Project description:Viperin is a radical SAM enzyme that has been shown to possess antiviral activity against a broad spectrum of viruses; however, its molecular mechanism is unknown. We report here that recombinant fungal and archaeal viperin enzymes catalyze the addition of the 5'-deoxyadenosyl radical (5'-dA•) to the double bond of isopentenyl pyrophosphate (IPP), producing a new compound we named adenylated isopentyl pyrophosphate (AIPP). The reaction is specific for IPP, as other pyrophosphate compounds involved in the mevalonate biosynthetic pathway did not react with 5'-dA• Enzymatic reactions employing IPP derivatives as substrates revealed that any chemical change in IPP diminishes its ability to be an effective substrate of fungal viperin. Mutational studies disclosed that the hydroxyl group on the side chain of Tyr-245 in fungal viperin is the likely source of hydrogen in the last step of the radical addition, providing mechanistic insight into the radical reaction catalyzed by fungal viperin. Structure-based molecular dynamics (MD) simulations of viperin interacting with IPP revealed a good fit of the isopentenyl motif of IPP to the active site cavity of viperin, unraveling the molecular basis of substrate specificity of viperin for IPP. Collectively, our findings indicate that IPP is an effective substrate of fungal and archaeal viperin enzymes and provide critical insights into the reaction mechanism.

| S-EPMC6130951 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data