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We report on the cDNA cloning and characterization of a novel short-chain isoprenyl diphosphate synthase from the aphid Myzus persicae. Of the three IPPS cDNAs we cloned, two yielded prenyltransferase activity following expression in Escherichia coli; these cDNAs encode identical proteins except for the presence, in one of them, of an N-terminal mitochondrial targeting peptide. Although the aphid enzyme was predicted to be a farnesyl diphosphate synthase by BLASTP analysis, rMpIPPS, when isopentenyl diphosphate and dimethylallyl diphosphate are supplied as substrates, typically generated geranyl diphosphate (C10) as its main product, along with significant quantities of farnesyl diphosphate (C15). Analysis of an MpIPPS homology model pointed to substitutions that could confer GPP/FPP synthase activity to the aphid enzyme.

ComX pheromone is an isoprenoidal oligopeptide containing a modified tryptophan residue, which stimulates natural genetic competence in the gram-positive bacterium Bacillus. Since posttranslational prenylation on the tryptophan residue has not been reported except in ComX pheromone, the universality of this modification has not yet been elucidated. In this paper, we established a cell-free system, whereby the tryptophan residue in peptides is modified with a geranyl group by modifying enzyme ComQ. In addition, we investigated enzymatic reaction conditions using an in vitro enzyme reaction system. This is the first report of in vitro geranylation on the tryptophan residue. This system is potentially a useful tool for elucidating the universality of prenylation on the tryptophan residue.

Various plant developmental processes involve phytohormones such as cytokinins. Isopentenyltransferase (IPT) reaction is the key rate-limiting step in cytokinin biosynthesis that transfers the isopentenyl (iP) group from dimethylallyl diphosphate to the N6-amino group of adenine. Here, a series of diadenosine polyphosphates (Ap(n)A) were screened as possible substrates of IPT, among which diadenosine tetraphosphate, diadenosine pentaphosphate and diadenosine hexaphosphate showed higher affinity than did the authentic substrates ADP and ATP. In addition, formation of mono-isopentenyl Ap(n)A and di-isopentenyl Ap(n)A was observed. Judging by the existing biosynthetic and hydrolytic systems for Ap(n)A in plants, Ap(n)A and isopentenyl-Ap(n)A may occur in the plant cells, with functional importance.

Faraesyl pyrophosphate synthase (FPP) displaying dimethylallyl transferase activity (EC 2.5.1.1) and geranyl transferase activity (EC 2.5.1.10) was purified from Capsicum fruits. This prenyltransferase has a molecular mass of 89000 ± 5000 Da resulting from the association of two apparently identical subunits having a molecular mass of 43000 ± 2000 Da. Antibodies raised against Capsicum FPP synthase selectively blocked the transferase activity. Analysis of the immunological relationships between FPP synthase and geranylgeranyl pyrophosphate synthase (EC 2.5.1.1, EC 2.5.1.10 and EC 2.5.1.30) revealed that these two enzymes though performing the same mechanism of catalysis and accepting identical substrates have different antigenic determinants. Thus, in connection to previous work, this immunological study suggests that Capsicum FPP is strictly located in the extraplastidial compartment.

Fnq26 from Streptomyces cinnamonensis DSM 1042 is a new member of the recently identified CloQ/Orf2 class of prenyltransferases. The enzyme was overexpressed in E. coli and purified to apparent homogeneity, resulting in a soluble, monomeric protein of 33.2kDa. The catalytic activity of Fnq26 is independent of the presence of Mg2+ or other divalent metal ions. With flaviolin (2,5,7-trihydroxy-1,4-naphthoquinone) as substrate, Fnq26 catalyzes the formation of a carbon–carbon-bond between C-3 (rather than C-1) of geranyl diphosphate and C-3 of flaviolin, i.e. an unusual “reverse” prenylation. With 1,3-dihydroxynaphthalene and 4-hydroxybenzoate as substrates Fnq26 catalyzes O-prenylations.

A new prenyltransferase (PT) enzyme derived from the microsomal fractions of cell cultures of Morus nigra was shown to be able to prenylate exclusively chalcones with a 2',4'-dihydroxy substitution and the isoflavone genistein. Computational studies were performed to shed some light on the relationship between the structure of the substrate and the enzymatic activity. PT requires divalent cations, particularly Mg(2+), to be effective. The apparent K(m) values for gamma,gamma-dimethylallyldiphosphate and 2',4'-dihydroxychalcone were 63 and 142 microM, respectively. The maximum activity of the enzyme was expressed during the first 10 days of cell growth.

The deduced polypeptide sequence of open reading frame slr1736 reveals homology to chlorophyll synthase and 1,4-dihydroxy-2-naphthoic acid phytyltransferase in Synechocystis sp. strain PCC 6803. In tocopherol and plastoquinone biosynthesis, a condensation reaction mechanistically similar to that of these two enzymes is performed. To analyze the function of this novel prenyltransferase, a deletion mutant of slr1736 was generated by homologous recombination. The mutant showed a markedly decreased tocopherol content, while plastoquinone levels remained unchanged. Since the aromatic precursor homogentisic acid accumulated in the mutant, the function of the enzyme was proven to be a novel tocopherol phytyltransferase.

Sulfolobus acidocaldarius geranylgeranyl diphosphate synthase yields (all-E)-C(20) prenyl diphosphate as a final product. The three-dimensional model of the enzyme suggested that removing two bulky residues at 77 and 114 would allow additional prenyl-chain elongation. To test this, we examined several mutants with substitutions at 77 and/or 114. As a result, the mutants, F77G, F77G and H114A, F77G and H114G, H114A, and H114G gave C(30), C(45), C(50), C(30) and C(40) as the main long product, respectively. These observations indicate that histidine-114 plays a crucial role in chain-length determination along with phenylalanine-77.

Two prenyltransferases were located in cell cultures of Vitis vinifera. A geranyl pyrophosphate synthase (EC 2.5.1.1) was associated with plastid-like membranes whereas a farnesyl pyrophosphate synthase (EC 2.5.1.10) was found to be soluble.

Heptaprenyl pyrophosphate synthetase from Bacillus subtilis was dissociated into two essential components, none of which had catalytic activity alone. The enzyme activity was restored when the two components were combined with each other. Both fractions, designated components I and II in the order of their elution from DEAE-Sephadex, appeared to be proteins of Mr 30 000. Component I was much more stable than component II which was easily destroyed by relatively mild heat treatment. Neither was inter-changeable with any of the essential components of hexaprenyl pyrophosphate synthetase of Micrococcus luteus B-P 26.

A cDNA of Chlamydomonas reinhardtii encoding a plastidial homogentisate prenyltransferase was identified. Functional expression studies in Escherichia coli revealed that the enzyme possessed properties similar to the prenyltransferase of Arabidopsis thaliana encoded by At3g11950 but different from the phytyltransferases of A. thaliana and Synechocystis. Unlike the phytyltransferases, the C. reinhardtii and the respective A. thaliana enzyme showed highest activities with solanesyl diphosphate, but were hardly active with phytyl diphosphate. Hence, these data provide evidence that the latter represent homogentisate solanesyltransferases involved in plastoquinone-9 biosynthesis. Overexpression of At3g11950 in A. thaliana, however, suggests that the solanesyltransferase can affect tocopherol biosynthesis as well.

A prenyltransferase (EC 2.5.1.1) was isolated and partially purified from cell cultures of Lithospermum erythrorhizon. The product of the enzymatic reaction is geranyl pyrophosphate rather than farnesyl pyrophosphate.

A microsomal fraction of elicitor-challenged bean (Phaseolus vulgaris) cell suspension cultures catalyses the prenylation of 3,9-dihydroxypterocarpan (DHP) to 3,9-dihydroxy-10-dimethylallylpterocarpan (phaseollidin) with dimethylallylpyrophosphate (DMAPP) as the prenyl donor. Km values for DMAPP and DHP are in the range 1–3 μM. Strong product inhibition with phaseollidin and phaseollin is observed.