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Dieter Strack

Publications and source records attributed to Dieter Strack.

33 records · Page 2Linked to original sources
PubMed2003

A flavonol O-methyltransferase from Catharanthus roseus performing two sequential methylations.

Protein extracts from dark-grown cell suspension cultures of Catharanthus roseus (Madagascar periwinkle) contained several O-methyltransferase (OMT) activities, including the 16-hydroxytabersonine O-methyltransferase (16HT-OMT) in indole alkaloid biosynthesis. This enzyme was enriched through several purification steps, including affinity chromatography on adenosine agarose. SDS-PAGE of the purified protein preparation revealed a protein band at the size expected for plant OMTs (38-43 kDa). Mass spectrometry indicated two dominant protein species of similar mass in this band, and sequences of tryptic peptides showed similarities to known OMTs. Homology-based RT-PCR identified cDNAs for four new OMTs. Two of these cDNAs (CrOMT2 and CrOMT4) encoded the proteins dominant in the preparation enriched for 16HT-OMT. The proteins were closely related (73% identity), but both shared only 48-53% identity with the closest relatives found in the public databases. The enzyme functions were investigated with purified recombinant proteins after cDNA expression in Escherichia coli. Unexpectedly, both proteins had no detectable 16HT-OMT activity, and CrOMT4 was inactive with all substrates investigated. CrOMT2 was identified as a flavonoid OMT that was expressed in dark-grown cell cultures and copurified with 16HT-OMT. It represented a new type of OMT that performs two sequential methylations at the 3'- and 5'-positions of the B-ring in myricetin (flavonol) and dihydromyricetin (dihydroflavonol). The resulting methylation pattern is characteristic for C. roseus flavonol glycosides and anthocyanins, and it is proposed that CrOMT2 is involved in their biosynthesis.

Amino Acid Sequence↗
PubMed2003

Antioxidant flavonoids from leaves of Polygonum hydropiper L.

Ten flavonoid compounds were isolated from the dried leaves of Polygonum hydropiper L. (Laksa leaves), and identified as 3-O-alpha-L-rhamnopyranosyloxy-3',4',5,7-tetrahydroxyflavone; 3-O-beta-D-glucopyranosyloxy-4',5,7-trihydroxyflavone; 6-hydroxyapigenin; 6"-O-(3,4,5-trihydroxybenzoyl) 3-O-beta-D-glucopyranosyloxy-3', 4', 5, 7-tetrahydroxyflavone; scutillarein; 6-hydroxyluteolin; 3',4',5,6,7-pentahydroxyflavone; 6-hydroxyluteolin-7-O-beta-D-glucopyranoside; quercetin 3-O-beta-D-glucuronide; 2"-O-(3,4,5-trihydroxybenzoyl) quercitrin; quercetin. Evaluation of the antioxidative activity, conducted in vitro, by using electron spin resonance (ESR) and ultraviolet visible (UV-vis) spectrophotometric assays, showed that these isolated flavonoids possess strong antioxidative capabilities. Measurement of the Trolox equivalent antioxidant capacity (TEAC) values, against ABTS (2,2'-azinobis(3-ethyl-benzo-thiazoline-6-sulphonic acid) radicals and phenyl-tert-butyl nitrone (PBN) azo initiator (AI) also showed strong anti-oxidative activity. The most powerful of the antioxidants was 2"-O-(3,4,5-trihydroxybenzoyl) quercitrin (galloyl quercitrin). A combination of two flavonoid compounds was tested for synergistic anti-oxidative capacity, but no significant improvement was observed.

Antioxidants↗
PubMed2003

Stilbenecarboxylate biosynthesis: a new function in the family of chalcone synthase-related proteins.

Chalcone (CHS), stilbene (STS) synthases, and related proteins are key enzymes in the biosynthesis of many secondary plant products. Precursor feeding studies and mechanistic rationalization suggest that stilbenecarboxylates might also be synthesized by plant type III polyketide synthases; however, the enzyme activity leading to retention of the carboxyl moiety in a stilbene backbone has not yet been demonstrated. Hydrangea macrophylla L. (Garden Hortensia) contains stilbenecarboxylates (hydrangeic acid and lunularic acid) that are derived from 4-coumaroyl and dihydro-4-coumaroyl starter residues, respectively. We used homology-based techniques to clone CHS-related sequences, and the enzyme functions were investigated with recombinant proteins. Sequences for two proteins were obtained. One was identified as CHS. The other shared 65-70% identity with CHSs and other family members. The purified recombinant protein had stilbenecarboxylate synthase (STCS) activity with dihydro-4-coumaroyl-CoA, but not with 4-coumaroyl-CoA or other substrates. We propose that the enzyme is involved in the biosynthesis of lunularic acid. It is the first example of a STS-type reaction that does not lose the terminal carboxyl group during the ring folding to the end product. Comparisons with CHS, STS, and a pyrone synthase showed that it is the only enzyme exerting a tight control over decarboxylation reactions. The protein contains unusual residues in positions highly conserved in other CHS-related proteins, and mutagenesis studies suggest that they are important for the structure or/and the catalytic activity. The formation of the natural products in vivo requires a reducing step, and we discuss the possibility that the absence of a reductase in the in vitro reactions may be responsible for the failure to obtain stilbenecarboxylates from substrates like 4-coumaroyl-CoA.

Acyltransferases↗
PubMed2003

Recent advances in betalain research.

Betalains replace the anthocyanins in flowers and fruits of plants of most families of the Caryophyllales. Unexpectedly, they were also found in some higher fungi. Whereas the anthocyanin-analogous functions of betalains in flower and fruit colouration are obvious, their role in fungi remains obscure. The nature of newly identified betalains as well as final structure elucidation of earlier putatively described compounds published within the last decade is compiled in this report. Recent advances in research on betalain biosynthesis is also covered, including description of some 'early' reactions, i.e. betalain-specific dopa formation in plants and fungi and extradiolic dopa cleavage in fungi. Work on betalain-specific glucosyltransferases (GTs) has given new insights into the evolution of secondary plant enzymes. It is proposed that these GTs are phylogenetically related to flavonoid GTs. It was found that the decisive steps in betalain biosynthesis, i.e. condensation of the betalain chromophore betalamic acid with cyclo-dopa and amino acids or amines in the respective aldimine formation of the red-violet betacyanins and the yellow betaxanthins, are most likely to be non-enzymatic. Betalains have attracted workers in applied fields because of their use for food colouring and their antioxidant and radical scavenging properties for protection against certain oxidative stress-related disorders.

Acyltransferases↗
PubMed2003

Arbuscular mycorrhiza: biological, chemical, and molecular aspects.

Mycorrhizas are the most important mutualistic symbioses on earth. The most prevalent type are the arbuscular mycorrhizas (AMs) that develop between roots of most terrestrial plants and fungal species of the Zygomycota. The AM fungi are able to grow into the root cortex forming intercellular hyphae from which highly branched structures, arbuscules, originate within cortex cells. The arbuscules are responsible for nutrient exchange between the host and the symbiont, transporting carbohydrates from the plant to the fungus and mineral nutrients, especially phosphate, and water from the fungus to the plant. Plants adapt their phosphate uptake to the interaction with the AM fungus by synthesis of specific phosphate transporters. Colonization of root cells induces dramatic changes in the cytoplasmic organization: vacuole fragmentation, transformation of the plasma membrane to a periarbuscular membrane covering the arbuscule, increase of the cytoplasm volume and numbers of cell organelles, as well as movement of the nucleus into a central position. The plastids form a dense network covering the symbiotic interface. In some of these changes, microtubules are most likely involved. With regard to the molecular crosstalk between the two organisms, a number of phytohormones (cytokinins, abscisic acid, jasmonate) as well as various secondary metabolites have been examined: (i) Jasmonates occur at elevated level, which is accompanied by cell-specific expression of genes involved in jasmonate biosynthesis that might be linked to strong carbohydrate sink function of AM roots and induced defense reactions: (ii) apocarotenoids (derivatives of mycorradicin and glycosylated cyclohexenones) accumulate in most mycorrhizal roots examined so far. Their biosynthesis via the nonmevalonate methylerythritol phosphate (MEP) pathway has been studied resulting in new insights into AM-specific gene expression and biosynthesis of secondary isoprenoids.

Carbohydrates↗
PubMed2002

Stimulation of carotenoid metabolism in arbuscular mycorrhizal roots.

Development of arbuscular mycorrhizal roots is correlated with accumulation of various isoprenoids, i.e. acyclic C(14) polyene 'mycorradicin' and C(13) cyclohexenone derivatives. We present data indicating a strong stimulation of carotenoid metabolism in such roots. Carotenoid profiling revealed mycorrhiza-specific accumulation of zeta-carotene in Zea mays and Medicago truncatula. Precursor accumulation after inhibition of phytoene desaturase (Pds) activity by norflurazon indicated an increased phytoene biosynthetic capacity in mycorrhizal roots of all species analyzed. Nicotiana tabacum plants transformed with a PDS promoter- GUS construct showed a cell-specific induction of PDS promoter activity in root cells containing arbuscules. Mycorradicin biosynthesis and, partially, mycorrhization were impaired in maize mutants deficient in carotenoid biosynthesis. These data indicate that (1) mycorradicin is probably synthesized via a C(40) precursor carotenoid, (2) carotenoid biosynthesis is induced in mycorrhizal roots, (3) induction occurs, at least partially, at the transcriptional level, and (4) that this may play a functional role during mycorrhization.

Carotenoids↗
PubMed2002

Immunolocalization of 1- O-sinapoylglucose:malate sinapoyltransferase in Arabidopsis thaliana.

The serine carboxypeptidase-like protein 1- O-sinapoylglucose:malate sinapoyltransferase (SMT) catalyzes the transfer of the sinapoyl moiety of 1- O-sinapoylglucose to malate in the formation of sinapoylmalate in some members of the Brassicaceae. Rabbit polyclonal monospecific antibodies were raised against the recombinant SMT produced in Escherichia coli from the corresponding Arabidopsis thaliana (L.) Heynh. cDNA. Immunoblot analysis of protein from different Arabidopsis tissues showed that the SMT is produced in all plant organs, except in the seeds and young seedlings. The enzyme was most abundant in older seedlings as well as in rosette leaves and the flowering stem of the plant. Minor amounts were found in the cauline leaves, flower buds and siliques. Traces were detected in the root and flowers. Arabidopsis and transgenic tobacco ( Nicotiana tabacum L.) plants expressing the full-length Arabidopsis SMT containing an N-terminal signal peptide showed apparent molecular masses of the protein of 52-55 kDa. The difference of ca. 8 kDa compared to the recombinant protein produced in E. coli was shown to be due to post-translational N-glycosylation of SMT in plants. Immunofluorescent labeling of Arabidopsis leaf sections localized SMT to the central vacuoles of mesophyll and epidermal cells. Comparable leaf sections of an SMT deletion mutant showed no vacuolar immunofluorescent labeling. We conclude that Arabidopsis SMT is synthesized as a precursor protein that is targeted to the endoplasmic reticulum where the signal peptide is removed. The correct N-terminus of the recombinantly produced SMT protein lacking the signal peptide was confirmed by Edman degradation. The protein is probably glycosylated in the Golgi apparatus from where it is subsequently routed to the vacuole.

Acyltransferases↗
PubMed2002

Accumulation of tyrosol glucoside in transgenic potato plants expressing a parsley tyrosine decarboxylase.

As part of the response to pathogen infection, potato plants accumulate soluble and cell wall-bound phenolics such as hydroxycinnamic acid tyramine amides. Since incorporation of these compounds into the cell wall leads to a fortified barrier against pathogens, raising the amounts of hydroxycinnamic acid tyramine amides might positively affect the resistance response. To this end, we set out to increase the amount of tyramine, one of the substrates of the hydroxycinnamoyl-CoA:tyramine N-(hydroxycinnamoyl)-transferase reaction, by placing a cDNA encoding a pathogen-induced tyrosine decarboxylase from parsley under the control of the 35S promoter and introducing the construct into potato plants via Agrobacterium tumefaciens-mediated transformation. While no alterations were observed in the pattern and quantity of cell wall-bound phenolic compounds in transgenic plants, the soluble fraction contained several new compounds. The major one was isolated and identified as tyrosol glucoside by liquid chromatography-electrospray ionization-high resolution mass spectrometry and NMR analyses. Our results indicate that expression of a tyrosine decarboxylase in potato does not channel tyramine into the hydroxycinnamoyl-CoA:tyramine N-(hydroxycinnamoyl)-transferase reaction but rather unexpectedly, into a different pathway leading to the formation of a potential storage compound.

Chromatography, High Pressure Liquid↗
PubMed2002

Two distantly related genes encoding 1-deoxy-d-xylulose 5-phosphate synthases: differential regulation in shoots and apocarotenoid-accumulating mycorrhizal roots.

Isopentenyl diphosphate, the universal precursor of isoprenoids, is synthesized by two separate routes, one in the cytosol and the other in plastids. The initial step of the plastidial pathway is catalysed by 1-deoxy-d-xylulose 5-phosphate synthase (DXS), which was previously thought to be encoded by a single-copy gene. We have identified two distinct classes of DXS-like cDNAs from the model legume Medicago truncatula. The deduced mature MtDXS1 and MtDXS2 proteins, excluding the predicted plastid-targeting peptides, are similar in size (72.7 and 71.2 kDa) yet share only 70% identity in their amino acid sequences, and both encode functional DXS proteins as shown by heterologous expression in Escherichia coli. Available DXS sequences from other plants can easily be assigned to either class 1 or class 2. Partial sequences of multiple DXS genes in a single genome may be found in the databases of several monocot and dicot plants. Blot analyses of RNA from M. truncatula, maize, tomato and tobacco demonstrate preferential expression of DXS1 genes in many developing plant tissues except roots. By contrast, DXS2 transcript levels are low in most tissues but are strongly stimulated in roots upon colonization by mycorrhizal fungi, correlated with accumulation of carotenoids and apocarotenoids. Monoterpene-synthesizing gland cells of leaf trichomes appear to be another site of DXS2 gene activity. The potential importance of DXS1 in many housekeeping functions and a still hypothetical role of DXS2 in the biosynthesis of secondary isoprenoids is discussed.

Amino Acid Sequence↗
PubMed2002

Occurrence and localization of apocarotenoids in arbuscular mycorrhizal plant roots.

The core structure of the yellow pigment from arbuscular mycorrhizal (AM) maize roots contains the apocarotenoids mycorradicin (an acyclic C14 polyene) and blumenol C cellobioside (a C13 cyclohexenone diglucoside). The pigment seems to be a mixture of different esterification products of these apocarotenoids. It is insoluble in water and accumulates as hydrophobic droplets in the vacuoles of root cortical cells. Screening 58 species from 36 different plant families, we detected mycorradicin in mycorrhizal roots of all Liliopsida analyzed and of a considerable number of Rosopsida, but also species were found in which mycorradicin was undetectable in mycorrhizal roots. Kinetic experiments and microscopic analyses indicate that accumulation of the yellow pigment is correlated with the concomitant degradation of arbuscules and the extensive plastid network covering these haustorium-like fungal structures. The role of the apocarotenoids in mycorrhizal roots is still unknown. The potential C40 carotenoid precursors, however, are more likely to be of functional importance in the development and functioning of arbuscules.

Carbohydrate Sequence↗
PubMed2002

Induction of jasmonate biosynthesis in arbuscular mycorrhizal barley roots.

Colonization of barley (Hordeum vulgare cv Salome) roots by an arbuscular mycorrhizal fungus, Glomus intraradices Schenck & Smith, leads to elevated levels of endogenous jasmonic acid (JA) and its amino acid conjugate JA-isoleucine, whereas the level of the JA precursor, oxophytodienoic acid, remains constant. The rise in jasmonates is accompanied by the expression of genes coding for an enzyme of JA biosynthesis (allene oxide synthase) and of a jasmonate-induced protein (JIP23). In situ hybridization and immunocytochemical analysis revealed that expression of these genes occurred cell specifically within arbuscule-containing root cortex cells. The concomitant gene expression indicates that jasmonates are generated and act within arbuscule-containing cells. By use of a near-synchronous mycorrhization, analysis of temporal expression patterns showed the occurrence of transcript accumulation 4 to 6 d after the appearance of the first arbuscules. This suggests that the endogenous rise in jasmonates might be related to the fully established symbiosis rather than to the recognition of interacting partners or to the onset of interaction. Because the plant supplies the fungus with carbohydrates, a model is proposed in which the induction of JA biosynthesis in colonized roots is linked to the stronger sink function of mycorrhizal roots compared with nonmycorrhizal roots.

Cyclopentanes↗
PubMed1998

Betanidin formation from dihydroxyphenylalanine in a model assay system.

Formation of betanidin, the aglycone of the red-violet betacyanins, has been demonstrated by a two-step model assay system. In the first step, dihydroxyphenylalanine (Dopa) was incubated with a Dopa dioxygenase preparation from Amanita muscaria, resulting in the formation of 4,5-seco-Dopa that spontaneously recyclized to betalamic acid. In the second step, a tyrosinase preparation from Portulaca grandiflora was added to the Dopa dioxygenase assay, resulting in Dopa oxidation followed by a spontaneous formation of cyclo-Dopa that, in turn, reacted spontaneously with betalamic acid to form betanidin. Thus, two enzymatic reactions, Dopa extradiol ring cleavage by the fungal enzyme and Dopa oxidation by the plant enzyme, initiate three spontaneous steps: the formation of cyclo-Dopa and betalamic acid and finally the condensation of these compounds to betanidin.

Journal Article↗
PubMedDate not supplied

Isolation and identification of arctiin and arctigenin in leaves of burdock (Arctium lappa L.) by polyamide column chromatography in combination with HPLC-ESI/MS.

A simple method involving polyamide column chromatography in combination with HPLC-PAD and HPLC-ESI/MS for isolating and identifying two kinds of lignans, arctiin and arctigenin, in the leaves of burdock (Arctium lappa L.) has been established. After extraction of burdock leaves with 80% methanol, the aqueous phase of crude extracts was partitioned between water and chloroform and the aqueous phase was fractionated on a polyamide glass column. The fraction, eluting with 100% methanol, was concentrated and gave a white precipitate at 4 degrees C from which two main compounds were purified by semi-preparative HPLC. In comparison with the UV and ESI-MS spectra and the HPLC retention time of authentic standards, the compounds were determined to be arctiin and arctigenin. The extraction/separation technique was validated using an internal standard method.

Arctium↗