03410nas a2200289   4500008004100000245014600041210006900187260000800256300001300264490000800277520240500285653002802690653002402718653002302742653002002765653001902785653002802804653002402832653001702856653002102873653001902894653001402913100002102927700002102948700002002969856013102989       2005                        eng d00aMonoterpene metabolism. Cloning, expression, and characterization of (-)-isopiperitenol/(-)-carveol dehydrogenase of peppermint and spearmint0 aMonoterpene metabolism Cloning expression and characterization o  cMar  a863–720 v1373 aThe essential oils of peppermint (Mentha x piperita) and spearmint (Mentha spicata) are distinguished by the oxygenation position on the p-menthane ring of the constitutive monoterpenes that is conferred by two regiospecific cytochrome P450 limonene-3- and limonene-6-hydroxylases. Following hydroxylation of limonene, an apparently similar dehydrogenase oxidizes (-)-trans-isopiperitenol to (-)-isopiperitenone in peppermint and (-)-trans-carveol to (-)-carvone in spearmint. Random sequencing of a peppermint oil gland secretory cell cDNA library revealed a large number of clones that specified redox-type enzymes, including dehydrogenases. Full-length dehydrogenase clones were screened by functional expression in Escherichia coli using a recently developed in situ assay. A single full-length acquisition encoding (-)-trans-isopiperitenol dehydrogenase (ISPD) was isolated. The (-)-ISPD cDNA has an open reading frame of 795 bp that encodes a 265-residue enzyme with a calculated molecular mass of 27,191. Nondegenerate primers were designed based on the (-)-trans-ISPD cDNA sequence and employed to screen a spearmint oil gland secretory cell cDNA library from which a 5'-truncated cDNA encoding the spearmint homolog, (-)-trans-carveol-dehydrogenase, was isolated. Reverse transcription-PCR amplification and RACE were used to acquire the remaining 5'-sequence from RNA isolated from oil gland secretory cells of spearmint leaf. The full-length spearmint dehydrogenase shares >99% amino acid identity with its peppermint homolog and both dehydrogenases are capable of utilizing (-)-trans-isopiperitenol and (-)-trans-carveol. These isopiperitenol/carveol dehydrogenases are members of the short-chain dehydrogenase/reductase superfamily and are related to other plant short-chain dehydrogenases/reductases involved in secondary metabolism (lignan biosynthesis), stress responses, and phytosteroid biosynthesis, but they are quite dissimilar (approximately 13% identity) to the monoterpene reductases of mint involved in (-)-menthol biosynthesis. The isolation of the genes specifying redox enzymes of monoterpene biosynthesis in mint indicates that these genes arose from different ancestors and not by simple duplication and differentiation of a common progenitor, as might have been anticipated based on the common reaction chemistry and structural similarity of the substrate monoterpenes.10aAlcohol Oxidoreductases10aAmino Acid Sequence10aCloning: Molecular10aMentha piperita10aMentha spicata10aMolecular Sequence Data10aMolecular Structure10aMonoterpenes10aMultigene Family10aOils: Volatile10aPhylogeny1 aRinger, Kerry, L1 aDavis, Edward, M1 aCroteau, Rodney  uhttps://lamiaceae.myspecies.info/content/monoterpene-metabolism-cloning-expression-and-characterization-isopiperitenol-carveol03113nas a2200457   4500008004100000245018300041210006900224260000800293300001300301490000700314520157800321653002401899653001301923653002301936653002101959653001201980653002501992653002002017653004402037653003802081653001602119653002302135653002802158653002402186653003102210653001302241653002102254653001402275653002502289653003402314653002602348100001902374700001502393700001902408700001502427700001802442700002202460700002002482700002002502856013302522       2002                        eng d00aCharacterization of phenylpropene O-methyltransferases from sweet basil: facile change of substrate specificity and convergent evolution within a plant O-methyltransferase family0 aCharacterization of phenylpropene Omethyltransferases from sweet  cFeb  a505–190 v143 aSome basil varieties are able to convert the phenylpropenes chavicol and eugenol to methylchavicol and methyleugenol, respectively. Chavicol O-methyltransferase (CVOMT) and eugenol O-methyltransferase (EOMT) cDNAs were isolated from the sweet basil variety EMX-1 using a biochemical genomics approach. These cDNAs encode proteins that are 90% identical to each other and very similar to several isoflavone O-methyltransferases such as IOMT, which catalyzes the 4'-O-methylation of 2,7,4'-trihydroxyisoflavanone. On the other hand, CVOMT1 and EOMT1 are related only distantly to (iso)eugenol OMT from Clarkia breweri, indicating that the eugenol O-methylating enzymes in basil and C. breweri evolved independently. Transcripts for CVOMT1 and EOMT1 were highly expressed in the peltate glandular trichomes on the surface of the young basil leaves. The CVOMT1 and EOMT1 cDNAs were expressed in Escherichia coli, and active proteins were produced. CVOMT1 catalyzed the O-methylation of chavicol, and EOMT1 also catalyzed the O-methylation of chavicol with equal efficiency to that of CVOMT1, but it was much more efficient in O-methylating eugenol. Molecular modeling, based on the crystal structure of IOMT, suggested that a single amino acid difference was responsible for the difference in substrate discrimination between CVOMT1 and EOMT1. This prediction was confirmed by site-directed mutagenesis, in which the appropriate mutants of CVOMT1 (F260S) and EOMT1 (S261F) were produced that exhibited the opposite substrate preference relative to the respective native enzyme.10aAmino Acid Sequence10aAnisoles10aDNA: Complementary10aEscherichia coli10aEugenol10aEvolution: Molecular10aGene Expression10aGene Expression Regulation: Enzymologic10aGene Expression Regulation: Plant10aIsoflavones10aMethyltransferases10aMolecular Sequence Data10aMolecular Structure10aMutagenesis: Site-Directed10aMutation10aOcimum basilicum10aPhylogeny10aRecombinant Proteins10aSequence Homology: Amino Acid10aSubstrate Specificity1 aGang, David, R1 aLavid, Noa1 aZubieta, Chloe1 aChen, Feng1 aBeuerle, Till1 aLewinsohn, Efraim1 aNoel, Joseph, P1 aPichersky, Eran  uhttps://lamiaceae.myspecies.info/content/characterization-phenylpropene-o-methyltransferases-sweet-basil-facile-change-substrate02657nas a2200325   4500008004100000245016800041210006900209260000800278300001400286490000800300520151200308653002401820653003801844653002201882653002801904653002401932653002101956653001401977653003401991653001302025653001302038100001702051700003202068700001802100700001902118700001502137700002202152700002002174856013702194       2004                        eng d00aThe biochemical and molecular basis for the divergent patterns in the biosynthesis of terpenes and phenylpropenes in the peltate glands of three cultivars of basil0 abiochemical and molecular basis for the divergent patterns in th  cNov  a3724–360 v1363 aSurface glandular trichomes distributed throughout the aerial parts of sweet basil (Ocimum basilicum) produce and store monoterpene, sesquiterpene, and phenylpropene volatiles. Three distinct basil chemotypes were used to examine the molecular mechanisms underlying the divergence in their monoterpene and sesquiterpene content. The relative levels of specific terpenes in the glandular trichomes of each cultivar were correlated with the levels of transcripts for eight genes encoding distinct terpene synthases. In a cultivar that produces mostly (R)-linalool, transcripts of (R)-linalool synthase (LIS) were the most abundant of these eight. In a cultivar that synthesizes mostly geraniol, transcripts of geraniol synthase were the most abundant, but the glands of this cultivar also contained a transcript of an (R)-LIS gene with a 1-base insertion that caused a frameshift mutation. A geraniol synthase-LIS hybrid gene was constructed and expressed in Escherichia coli, and the protein catalyzed the formation of both geraniol and (R)-linalool from geranyl diphosphate. The total amounts of terpenes were correlated with total levels of terpene synthase activities, and negatively correlated with levels of phenylpropanoids and phenylalanine ammonia lyase activity. The relative levels of geranyl diphosphate synthase and farnesyl diphosphate synthase activities did not correlate with the total amount of terpenes produced, but showed some correlation with the ratio of monoterpenes to sesquiterpenes.10aAmino Acid Sequence10aGene Expression Regulation: Plant10aGenetic Variation10aMolecular Sequence Data10aMolecular Structure10aOcimum basilicum10aPhylogeny10aSequence Homology: Amino Acid10aStyrenes10aTerpenes1 aIijima, Yoko1 aDavidovich-Rikanati, Rachel1 aFridman, Eyal1 aGang, David, R1 aBar, Einat1 aLewinsohn, Efraim1 aPichersky, Eran  uhttps://lamiaceae.myspecies.info/content/biochemical-and-molecular-basis-divergent-patterns-biosynthesis-terpenes-and-phenylpropenes01628nas a2200277   4500008004100000245006700041210006700108260000800175300001300183490000700196520077100203653001300974653001300987653001401000653002401014653004501038653001301083653001401096653001701110100002401127700002201151700002601173700002701199700002101226856010301247       2003                        eng d00aBenzoxazinoids and iridoid glucosides from four Lamium species0 aBenzoxazinoids and iridoid glucosides from four Lamium species  cDec  a1413–70 v643 aA new class of compounds for the plant family Lamiaceae, benzoxazinoids, was found in Lamium galeobdolon. From the aerial parts of the species were isolated the new 2-O-beta-D-glucopyranosyl-6-hydroxy-2H-1,4-benzoxazin-3(4H)-one (6-hydroxy blepharin) together with four known benzoxazinoids, DHBOA-Glc, blepharin, DIBOA, DIBOA-Glc, as well as harpagide, 8-O-acetyl-harpagide and salidroside. Eight known iridoid glucosides, 24-epi-pterosterone and verbascoside were isolated from Lamium amplexicaule, L. purpureum and L. garganicum. The iridoids, 5-deoxylamiol and sesamoside, as well as the phytoecdysone, 24-epi-pterosterone, were found for the first time for the genus Lamium. The phytochemical data are discussed from a systematic and evolutionary point of view.10aEcdysone10aIridoids10aLamiaceae10aMolecular Structure10aNuclear Magnetic Resonance: Biomolecular10aOxazines10aPhylogeny10aPhytosterols1 aAlipieva, Kalina, I1 aTaskova, Rilka, M1 aEvstatieva, Ljubka, N1 aHandjieva, Nedjalka, V1 aPopov, Simeon, S  uhttps://lamiaceae.myspecies.info/content/benzoxazinoids-and-iridoid-glucosides-four-lamium-species03120nas a2200349   4500008004100000245018100041210006900222260000800291300001400299490000800313520197500321653002102296653002402317653001702341653002802358653002402386653001402410653001702424653002102441653001102462100001402473700001602487700002702503700001302530700001602543700001302559700001402572700002002586700001402606700001502620856013502635       2001                        eng d00aMalonyl-CoA:anthocyanin 5-O-glucoside-6"'-O-malonyltransferase from scarlet sage (Salvia splendens) flowers. Enzyme purification, gene cloning, expression, and characterization0 aMalonylCoAanthocyanin 5Oglucoside6Omalonyltransferase from scarl  cDec  a49013–90 v2763 aThe orange to blue coloration of flowers in nature is, in most cases, provided by anthocyanins, a class of plant flavonoids, many of which are modified by malonyl group(s). However, the identity of the enzyme catalyzing the malonylation reaction remains to be established. Here, we describe for the first time the purification, characterization, and cDNA cloning of an anthocyanin malonyltransferase from scarlet sage (Salvia splendens) flowers. The purified enzyme (termed Ss5MaT1) was a monomeric 50-kDa protein catalyzing the regiospecific transfer of the malonyl group from malonyl-CoA to the 6"'-hydroxyl group of the 5-glucosyl moiety of anthocyanins. Ss5MaT1 showed a k(cat) value of 7.8 s(-1) at 30 degrees C and pH 7.0 for the malonylation of bisdemalonylsalvianin (pelargonidin 3-(6"-O-caffeyl-beta-glucopyranoside)-5-beta-glucopyranoside) and K(m) values of 101 microm and 57 microm for bisdemalonylsalvianin and malonyl-CoA, respectively. p-Coumaric acid, which mimics an aromatic acyl group linked to the 3-glucosidic moiety of an anthocyanin substrate, was a competitive inhibitor with respect to the substrate. This strongly suggests that the presence of an aromatic acyl group at the 3-glucosidic moiety of anthocyanin is important for substrate recognition by the enzyme. On the basis of the partial amino acid sequences of the purified enzyme, we isolated a cDNA encoding Ss5MaT1. Ss5MaT1 consisted of 462 amino acids and shared motifs that are commonly found among members of a versatile plant acyltransferase family, which was recently shown to include numerous homologs of unknown biochemical functions. Northern blot analysis revealed that the transcripts of Ss5MaT1 were detected in petals, sepals, bracts, and red stems, in accordance with the pigment accumulation patterns. Phylogenetic analysis suggests that the aliphatic and aromatic acylations of anthocyanins are generally catalyzed by subfamily members of the plant acyltransferase family.10aAcyltransferases10aAmino Acid Sequence10aAnthocyanins10aMolecular Sequence Data10aMolecular Structure10aPhylogeny10aPigmentation10aPlant Structures10aSalvia1 aSuzuki, H1 aNakayama, T1 aYonekura-Sakakibara, K1 aFukui, Y1 aNakamura, N1 aNakao, M1 aTanaka, Y1 aYamaguchi, M, A1 aKusumi, T1 aNishino, T  uhttps://lamiaceae.myspecies.info/content/malonyl-coaanthocyanin-5-o-glucoside-6-o-malonyltransferase-scarlet-sage-salvia-splendens02990nas a2200445   4500008004100000245012100041210006900162260000800231300001400239490000700253520151700260653002401777653002301801653001501824653002501839653002801864653004101892653003001933653001301963653001601976653002301992653002202015653002802037653002402065653002102089653001402110653001902124653002102143653001502164653005202179653003402231653002602265100002002291700002402311700001802335700001802353700002202371700001902393856013202412       2007                        eng d00aEvolution of Cinnamate/p-coumarate carboxyl methyltransferases and their role in the biosynthesis of methylcinnamate0 aEvolution of Cinnamatepcoumarate carboxyl methyltransferases and  cOct  a3212–290 v193 aMethylcinnamate, which is widely distributed throughout the plant kingdom, is a significant component of many floral scents and an important signaling molecule between plants and insects. Comparison of an EST database obtained from the glandular trichomes of a basil (Ocimum basilicum) variety that produces high levels of methylcinnamate (line MC) with other varieties producing little or no methylcinnamate identified several very closely related genes belonging to the SABATH family of carboxyl methyltransferases that are highly and almost exclusively expressed in line MC. Biochemical characterization of the corresponding recombinant proteins showed that cinnamate and p-coumarate are their best substrates for methylation, thus designating these enzymes as cinnamate/p-coumarate carboxyl methyltransferases (CCMTs). Gene expression, enzyme activity, protein profiling, and metabolite content analyses demonstrated that CCMTs are responsible for the formation of methylcinnamate in sweet basil. A phylogenetic analysis of the entire SABATH family placed these CCMTs into a clade that includes indole-3-acetic acid carboxyl methyltransferases and a large number of uncharacterized carboxyl methyltransferase-like proteins from monocots and lower plants. Structural modeling and ligand docking suggested active site residues that appear to contribute to the substrate preference of CCMTs relative to other members of the SABATH family. Site-directed mutagenesis of specific residues confirmed these findings.10aAmino Acid Sequence10aBlotting: Southern10aCinnamates10aEvolution: Molecular10aExpressed Sequence Tags10aGas Chromatography-Mass Spectrometry10aGene Expression Profiling10aKinetics10aMethylation10aMethyltransferases10aModels: Molecular10aMolecular Sequence Data10aMolecular Structure10aOcimum basilicum10aPhylogeny10aPlant Proteins10aProtein Isoforms10aProteomics10aReverse Transcriptase Polymerase Chain Reaction10aSequence Homology: Amino Acid10aSubstrate Specificity1 aKapteyn, Jeremy1 aQualley, Anthony, V1 aXie, Zhengzhi1 aFridman, Eyal1 aDudareva, Natalia1 aGang, David, R  uhttps://lamiaceae.myspecies.info/content/evolution-cinnamatep-coumarate-carboxyl-methyltransferases-and-their-role-biosynthesis