Geschmack ist vorhersagbar: Mit FlavorMiner. FlavorMiner heißt das Tool, das IPB-Chemiker und Partner aus Kolumbien jüngst entwickelt haben. Das Programm kann, basierend auf maschinellem Lernen (KI), anhand der…
Seit Februar 2021 bietet Wolfgang Brandt, ehemaliger Leiter der Arbeitsgruppe Computerchemie am IPB, sein Citizen Science-Projekt zur Pilzbestimmung an. Dafür hat er in regelmäßigen Abständen öffentliche Vorträge zur Vielfalt…
Hans, J.; Brandt, W.; Vogt, T.;Site-directed mutagenesis and protein 3D-homology modelling suggest a catalytic mechanism for UDP-glucose-dependent betanidin 5-O-glucosyltransferase from Dorotheanthus bellidiformisPlant J.39319-333(2004)DOI: 10.1111/j.1365-313X.2004.02133.x
In livingstone daisy (Dorotheanthus bellidiformis ), betanidin 5‐O‐glucosyltransferase (UGT73A5) is involved in the regiospecific glucosylation of betanidin and various flavonols. Based on sequence alignments several amino acid candidates which might be essential for catalysis were identified. The selected amino acids of the functionally expressed protein, suggested to be involved in substrate binding and turnover, were substituted via site‐directed mutagenesis. The substitution of two highly conserved amino acids, Glu378, located in the proposed UDP‐glucose binding site, and His22, located close to the N‐terminus, led to the complete loss of enzyme activity. A 3D model of this regiospecific betanidin and flavonoid glucosyltransferase was constructed and the active site modelled. This model was based on the crystallographic structure of a bacterial UDP‐glucose‐dependent glucosyltransferase from Amycolatopsis orientalis used as a template and the generated null mutations. To explain the observed inversion in the configuration of the bound sugar, semiempirical calculations favour an SN‐1 reaction, as one plausible alternative to the generally proposed SN‐2 mechanism discussed for plant natural product glucosyltransferases. The calculated structural data do not only explain the abstraction of a proton from the acceptor betanidin, but further imply that the reaction mechanism might also involve a catalytic triad, with similarities described for the serine protease family.
Publikation
Hans, J.; Hause, B.; Strack, D.; Walter, M. H.;Cloning, Characterization, and Immunolocalization of a Mycorrhiza-Inducible 1-Deoxy-D-Xylulose 5-Phosphate Reductoisomerase in Arbuscule-Containing Cells of MaizePlant Physiol.134614-624(2004)DOI: 10.1104/pp.103.032342
Colonization of plant roots by symbiotic arbuscular mycorrhizal fungi frequently leads to the accumulation of several apocarotenoids. The corresponding carotenoid precursors originate from the plastidial 2-C-methyl-d-erythritol 4-phosphate pathway. We have cloned and characterized 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR), catalyzing the first committed step of the pathway, from maize (Zea mays). Functional identification was accomplished by heterologous expression of sequences coding for the mature protein in Escherichia coli. DXR is up-regulated in maize roots during mycorrhization as shown at transcript and protein levels, but is also abundant in leaves and young seedlings. Inspection of sequenced genomes and expressed sequence tag (EST) databases argue for a single-copy DXR gene. Immunolocalization studies in mycorrhizal roots using affinity-purified antibodies revealed a DXR localization in plastids around the main symbiotic structures, the arbuscules. DXR protein accumulation is tightly correlated with arbuscule development. The highest level of DXR protein is reached around maturity and initial senescence of these structures. We further demonstrate the formation of a DXR-containing plastidial network around arbuscules, which is highly interconnected in the mature, functional state of the arbuscules. Our findings imply a functional role of a still unknown nature for the apocarotenoids or their respective carotenoid precursors in the arbuscular life cycle.
Publikation
Halitschke, R.; Ziegler, J.; Keinänen, M.; Baldwin, I. T.;Silencing of hydroperoxide lyase and allene oxide synthase reveals substrate and defense signaling crosstalk in Nicotiana attenuataPlant J.4035-46(2004)DOI: 10.1111/j.1365-313X.2004.02185.x
The fatty acid hydroperoxide (HP) substrates required for the biosynthesis of jasmonic acid (JA) and green leaf volatiles (GLVs) are supplied by separate lipoxygenases (LOX). We silenced the expression of two genes downstream of the LOX: allene oxide synthase (AOS) and HP lyase (HPL) by antisense expression of endogenous genes (NaAOS , NaHPL ) in Nicotiana attenuata , in which the biosynthesis of JA is amplified by herbivore‐specific elicitors. We report that these elicitors also amplify wound‐induced GLV releases, but suppress the wound‐induced increase of NaHPL transcripts, suggesting that substrate flux controls GLV biosynthesis. As expected, silencing of NaHPL and NaAOS reduced GLV release and JA accumulation, respectively. Surprisingly, HPL‐ and AOS‐silenced plants had enhanced JA and GLV responses, suggesting substrate ‘crosstalk’ between these two oxylipin cascades. Plants with depleted GLVs (as‐hpl ) were less attractive than wild type (WT) or empty vector control plants in choice‐tests with native lepidopteran herbivores. In feeding trials, Manduca sexta larvae developed slower on as‐hpl plants. The reduced larval consumption and performance, which was not caused by increases in defense responses in as‐hpl plants, could be restored to WT levels by the addition of synthetic GLVs, demonstrating that GLVs function as feeding stimulants. Gene expression profiling by cDNA microarray analysis and characterization of several induced defenses in herbivore‐elicited as‐hpl and as‐aos plants revealed differential involvement of JA and GLVs in defense signaling. Elicitation of volatile terpenoids (an indirect defense) requires JA signaling, where as trypsin protease inhibitor elicitation (a direct defense) requires both functional JA and GLV cascades.
The Phytophthora-derived oligopeptide elicitor, Pep-13, originally identified as an inducer of plant defense in the nonhost–pathogen interaction of parsley and Phytophthora sojae, triggers defense responses in potato. In cultured potato cells, Pep-13 treatment results in an oxidative burst and activation of defense genes. Infiltration of Pep-13 into leaves of potato plants induces the accumulation of hydrogen peroxide, defense gene expression and the accumulation of jasmonic and salicylic acids. Derivatives of Pep-13 show similar elicitor activity in parsley and potato, suggesting a receptor-mediated induction of defense response in potato similar to that observed in parsley. However, unlike in parsley, infiltration of Pep-13 into leaves leads to the development of hypersensitive response-like cell death in potato. Interestingly, Pep-13-induced necrosis formation, hydrogen peroxide formation and accumulation of jasmonic acid, but not activation of a subset of defense genes, is dependent on salicylic acid, as shown by infiltration of Pep-13 into leaves of potato plants unable to accumulate salicylic acid. Thus, in a host plant of Phytophthora infestans, Pep-13 is able to elicit salicylic acid-dependent and -independent defense responses.