zur Suche springenzur Navigation springenzum Inhalt springen

Sortieren nach: Erscheinungsjahr Typ der Publikation

Zeige Ergebnisse 1 bis 10 von 15.

Publikation

Reisberg, M.; Arnold, N.; Porzel, A.; Neubert, R. H.; Dräger, B.; Malusides, novel glucosylceramides isolated from apple pomace (Malus domestica) Z. Naturforsch. C 73, 33-39, (2018) DOI: 10.1515/znc-2017-0059

Three new glucosylceramides (GluCers) named malusides I–III (1–3) were isolated from apple (cultivars of Malus domestica) pomace (fruit material remaining after juice extraction). An unusual oxo/hydroxy group pattern within the sphingadienine (d18:2) type sphingoid base was observed. All compounds contained the same α-hydroxylated fatty acid (h16:0) and a β-D-glucose moiety. Their structures were assigned on the basis of one- and two-dimensional (1D and 2D) nuclear magnetic resonance (NMR) spectroscopic analyses and mass spectrometry (MS) measurements.
Publikation

Reisberg, M.; Arnold, N.; Bisrat, D.; Asres, K.; Neubert, R. H.; Dräger, B.; Quantification of glycosylceramides in plants by automated multiple development–high-performance thin-layer chromatography JPC - J. Planar Chromat. 30, 460-466, (2017) DOI: 10.1556/1006.2017.30.6.1

Glycosylceramides (GlyCers) are precursors of ceramides (Cers) that are major components of the outer layer of human skin, the stratum corneum. A Cer deficiency is associated with skin diseases such as psoriasis and atopic dermatitis and can be treated with Cer-containing semisolid formulations. Plants may serve as alternative sources for expensive semisynthetic Cer production. Since the GlyCer contents of plants vary widely, there is a need to develop a rapid, simple, selective, and precise method for GlyCer quantification in plants. In the present study, an effective and validated automated multiple development‒high-performance thin-layer chromatography (AMD‒HPTLC) method has been developed for GlyCer quantification in 9 different plant materials. An 18-step gradient elution program (n-hexane, chloroform, ethyl acetate, methanol) led to a clear separation of bands from complex matrices and allowed densitometric analysis for quantification purposes. Apple pomace and wheat germs yielded 26.8 and 39.5 mg of GlyCer per 100 g plant material, respectively, while the yields of coffee grounds were below the limit of quantification. The GlyCer contents of the seeds of six Fabaceae species, namely, Albizia grandibracteata, Albizia gummifera, Albizia lebbeck, Albizia schimperiana, Acacia etbaica, and Robinia pseudoacacia, ranged from 9.4 to 23.1 mg per 100 g plant material. GlyCers were separated by preparative thin-layer chromatography (TLC) and identified by offline high-performance liquid chromatography–mass spectrometry (HPLC–MS). Intact GlyCers were detected in the Fabaceae species for the first time. A simple AMD–HPTLC screening and quantification technique for GlyCers was developed, which may serve as a tool in searching plant GlyCers for a possible “phyto”-Cer production.
Publikation

Reisberg, M.; Arnold, N.; Porzel, A.; Neubert, R. H. H.; Dräger, B.; Production of Rare Phyto-Ceramides from Abundant Food Plant Residues J. Agr. Food Chem. 65, 1507-1517, (2017) DOI: 10.1021/acs.jafc.6b04275

Ceramides (Cers) are major components of the outermost layer of the skin, the stratum corneum, and play a crucial role in permeability barrier functions. Alterations in Cer composition causing skin diseases are compensated with semisynthetic skin-identical Cers. Plants constitute new resources for Cer production as they contain glucosylceramides (GluCers) as major components. GluCers were purified from industrial waste plant materials, apple pomace (Malus domestica), wheat germs (Triticum sp.), and coffee grounds (Coffea sp.), with GluCer contents of 28.9 mg, 33.7 mg, and 4.4 mg per 100 g of plant material. Forty-five species of GluCers (1–45) were identified with different sphingoid bases, saturated or monounsaturated α-hydroxy fatty acids (C15–28), and β-glucose as polar headgroup. Three main GluCers were hydrolyzed by a recombinant human glucocerebrosidase to produce phyto-Cers (46–48). These studies showed that rare and expensive phyto-Cers can be obtained from industrial food plant residues.
Publikation

Küster, N.; Rosahl, S.; Dräger, B.; Potato plants with genetically engineered tropane alkaloid precursors Planta 245, 355-365, (2017) DOI: 10.1007/s00425-016-2610-7

Main conclusionSolanum tuberosum tropinone reductase I reduced tropinone in vivo. Suppression of tropinone reductase II strongly reduced calystegines in sprouts. Overexpression of putrescine N -methyltransferase did not alter calystegine accumulation.Calystegines are hydroxylated alkaloids formed by the tropane alkaloid pathway. They accumulate in potato (Solanum tuberosum L., Solanaceae) roots and sprouting tubers. Calystegines inhibit various glycosidases in vitro due to their sugar-mimic structure, but functions of calystegines in plants are not understood. Enzymes participating in or competing with calystegine biosynthesis, including putrescine N-methyltransferase (PMT) and tropinone reductases (TRI and TRII), were altered in their activity in potato plants by RNA interference (RNAi) and by overexpression. The genetically altered potato plants were investigated for the accumulation of calystegines and for intermediates of their biosynthesis. An increase in N-methylputrescine provided by DsPMT expression was not sufficient to increase calystegine accumulation. Overexpression and gene knockdown of StTRI proved that S. tuberosum TRI is a functional tropinone reductase in vivo, but no influence on calystegine accumulation was observed. When StTRII expression was suppressed by RNAi, calystegine formation was severely compromised in the transformed plants. Under phytochamber and green house conditions, the StTRII RNAi plants did not show phenotypic alterations. Further investigation of calystegines function in potato plants under natural conditions is enabled by the calystegine deprived StTRII RNAi plants.
Publikation

Dobritzsch, M.; Lübken, T.; Eschen-Lippold, L.; Gorzolka, K.; Blum, E.; Matern, A.; Marillonnet, S.; Böttcher, C.; Dräger, B.; Rosahl, S.; MATE Transporter-Dependent Export of Hydroxycinnamic Acid Amides Plant Cell 28, 583-596, (2016) DOI: 10.1105/tpc.15.00706

The ability of Arabidopsis thaliana to successfully prevent colonization by Phytophthora infestans, the causal agent of late blight disease of potato (Solanum tuberosum), depends on multilayered defense responses. To address the role of surface-localized secondary metabolites for entry control, droplets of a P. infestans zoospore suspension, incubated on Arabidopsis leaves, were subjected to untargeted metabolite profiling. The hydroxycinnamic acid amide coumaroylagmatine was among the metabolites secreted into the inoculum. In vitro assays revealed an inhibitory activity of coumaroylagmatine on P. infestans spore germination. Mutant analyses suggested a requirement of the p-coumaroyl-CoA:agmatine N4-p-coumaroyl transferase ACT for the biosynthesis and of the MATE transporter DTX18 for the extracellular accumulation of coumaroylagmatine. The host plant potato is not able to efficiently secrete coumaroylagmatine. This inability is overcome in transgenic potato plants expressing the two Arabidopsis genes ACT and DTX18. These plants secrete agmatine and putrescine conjugates to high levels, indicating that DTX18 is a hydroxycinnamic acid amide transporter with a distinct specificity. The export of hydroxycinnamic acid amides correlates with a decreased ability of P. infestans spores to germinate, suggesting a contribution of secreted antimicrobial compounds to pathogen defense at the leaf surface.
Publikation

Reinhardt, N.; Fischer, J.; Coppi, R.; Blum, E.; Brandt, W.; Dräger, B.; Substrate flexibility and reaction specificity of tropinone reductase-like short-chain dehydrogenases Bioorg. Chem. 53, 37-49, (2014) DOI: 10.1016/j.bioorg.2014.01.004

Annotations of protein or gene sequences from large scale sequencing projects are based on protein size, characteristic binding motifs, and conserved catalytic amino acids, but biochemical functions are often uncertain. In the large family of short-chain dehydrogenases/reductases (SDRs), functional predictions often fail. Putative tropinone reductases, named tropinone reductase-like (TRL), are SDRs annotated in many genomes of organisms that do not contain tropane alkaloids. SDRs in vitro often accept several substrates complicating functional assignments. Cochlearia officinalis, a Brassicaceae, contains tropane alkaloids, in contrast to the closely related Arabidopsis thaliana. TRLs from Arabidopsis and the tropinone reductase isolated from Cochlearia (CoTR) were investigated for their catalytic capacity. In contrast to CoTR, none of the Arabidopsis TRLs reduced tropinone in vitro. NAD(H) and NADP(H) preferences were relaxed in two TRLs, and protein homology models revealed flexibility of amino acid residues in the active site allowing binding of both cofactors. TRLs reduced various carbonyl compounds, among them terpene ketones. The reduction was stereospecific for most of TRLs investigated, and the corresponding terpene alcohol oxidation was stereoselective. Carbonyl compounds that were identified to serve as substrates were applied for modeling pharmacophores of each TRL. A database of commercially available compounds was screened using the pharmacophores. Compounds identified as potential substrates were confirmed by turnover in vitro. Thus pharmacophores may contribute to better predictability of biochemical functions of SDR enzymes.
Publikation

Junker, A.; Fischer, J.; Sichhart, Y.; Brandt, W.; Dräger, B.; Evolution of the key alkaloid enzyme putrescine N-methyltransferase from spermidine synthase Front. Plant Sci. 4, 260, (2013) DOI: 10.3389/fpls.2013.00260

Putrescine N-methyltransferases (PMTs) are the first specific enzymes of the biosynthesis of nicotine and tropane alkaloids. PMTs transfer a methyl group onto the diamine putrescine from S-adenosyl-l-methionine (SAM) as coenzyme. PMT proteins have presumably evolved from spermidine synthases (SPDSs), which are ubiquitous enzymes of polyamine metabolism. SPDSs use decarboxylated SAM as coenzyme to transfer an aminopropyl group onto putrescine. In an attempt to identify possible and necessary steps in the evolution of PMT from SPDS, homology based modeling of Datura stramonium SPDS1 and PMT was employed to gain deeper insight in the preferred binding positions and conformations of the substrate and the alternative coenzymes. Based on predictions of amino acids responsible for the change of enzyme specificities, sites of mutagenesis were derived. PMT activity was generated in D. stramonium SPDS1 after few amino acid exchanges. Concordantly, Arabidopsis thaliana SPDS1 was mutated and yielded enzymes with both, PMT and SPDS activities. Kinetic parameters were measured for enzymatic characterization. The switch from aminopropyl to methyl transfer depends on conformational changes of the methionine part of the coenzyme in the binding cavity of the enzyme. The rapid generation of PMT activity in SPDS proteins and the wide-spread occurrence of putative products of N-methylputrescine suggest that PMT activity is present frequently in the plant kingdom.
Publikation

Jocković, N.; Fischer, W.; Brandsch, M.; Brandt, W.; Dräger, B.; Inhibition of Human Intestinal α-Glucosidases by Calystegines J. Agr. Food Chem. 61, 5550-5557, (2013) DOI: 10.1021/jf4010737

Calystegines are polyhydroxylated nortropane alkaloids found in Convolvulaceae, Solanaceae, and other plant families. These plants produce common fruits and vegetables. The calystegine structures resemble sugars and suggest interaction with enzymes of carbohydrate metabolism. Maltase and sucrase are α-glucosidases contributing to human carbohydrate degradation in the small intestine. Inhibition of these enzymes by orally administered drugs is one option for treatment of diabetes mellitus type 2. In this study, inhibition of maltase and sucrase by calystegines A3 and B2 purified from potatoes was investigated. In silico docking studies confirmed binding of both calystegines to the active sites of the enzymes. Calystegine A3 showed low in vitro enzyme inhibition; calystegine B2 inhibited mainly sucrose activity. Both compounds were not transported by Caco-2 cells indicating low systemic availability. Vegetables rich in calystegine B2 should be further investigated as possible components of a diet preventing a steep increase in blood glucose after a carbohydrate-rich meal.
Publikation

Biastoff, S.; Reinhardt, N.; Reva, V.; Brandt, W.; Dräger, B.; Evolution of putrescine N-methyltransferase from spermidine synthase demanded alterations in substrate binding FEBS Lett. 583, 3367-3374, (2009) DOI: 10.1016/j.febslet.2009.09.043

Putrescine N ‐methyltransferase (PMT) catalyses S ‐adenosylmethionine (SAM)‐dependent methylation of putrescine in tropane alkaloid biosynthesis. PMT presumably evolved from the ubiquitous spermidine synthase (SPDS). SPDS protein structure suggested that only few amino acid exchanges in the active site were necessary to achieve PMT activity. Protein modelling, mutagenesis, and chimeric protein construction were applied to trace back evolution of PMT activity from SPDS. Ten amino acid exchanges in Datura stramonium SPDS dismissed the hypothesis of facile generation of PMT activity in existing SPDS proteins. Chimeric PMT and SPDS enzymes were active and indicated the necessity for a different putrescine binding site when PMT developed.
Publikation

Biastoff, S.; Brandt, W.; Dräger, B.; Putrescine N-methyltransferase – The start for alkaloids Phytochemistry 70, 1708-1718, (2009) DOI: 10.1016/j.phytochem.2009.06.012

Putrescine N-methyltransferase (PMT) catalyses S-adenosylmethionine (SAM) dependent methylation of the diamine putrescine. The product N-methylputrescine is the first specific metabolite on the route to nicotine, tropane, and nortropane alkaloids. PMT cDNA sequences were cloned from tobacco species and other Solanaceae, also from nortropane-forming Convolvulaceae and enzyme proteins were synthesised in Escherichia coli. PMT activity was measured by HPLC separation of polyamine derivatives and by an enzyme-coupled colorimetric assay using S-adenosylhomocysteine. PMT cDNA sequences resemble those of plant spermidine synthases (putrescine aminopropyltransferases) and display little similarity to other plant methyltransferases. PMT is likely to have evolved from the ubiquitous enzyme spermidine synthase. PMT and spermidine synthase proteins share the same overall protein structure; they bind the same substrate putrescine and similar co-substrates, SAM and decarboxylated S-adenosylmethionine. The active sites of both proteins, however, were shaped differentially in the course of evolution. Phylogenetic analysis of both enzyme groups from plants revealed a deep bifurcation and confirmed an early descent of PMT from spermidine synthase in the course of angiosperm development.
IPB Mainnav Search