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…
Zurbriggen, M. D.; Carrillo, N.; Tognetti, V. B.; Melzer, M.; Peisker, M.; Hause, B.; Hajirezaei, M.-R.;Chloroplast-generated reactive oxygen species play a major role in localized cell death during the non-host interaction between tobacco and Xanthomonas campestris pv. vesicatoriaPlant J.60962-973(2009)DOI: 10.1111/j.1365-313X.2009.04010.x
Attempted infection of plants by pathogens elicits a complex defensive response. In many non‐host and incompatible host interactions it includes the induction of defence‐associated genes and a form of localized cell death (LCD), purportedly designed to restrict pathogen advance, collectively known as the hypersensitive response (HR). It is preceded by an oxidative burst, generating reactive oxygen species (ROS) that are proposed to cue subsequent deployment of the HR, although neither the origin nor the precise role played by ROS in the execution of this response are completely understood. We used tobacco plants expressing cyanobacterial flavodoxin to address these questions. Flavodoxin is an electron shuttle present in prokaryotes and algae that, when expressed in chloroplasts, specifically prevents ROS formation in plastids during abiotic stress episodes. Infiltration of tobacco wild‐type leaves with high titres of Xanthomonas campestris pv. vesicatoria (Xcv ), a non‐host pathogen, resulted in ROS accumulation in chloroplasts, followed by the appearance of localized lesions typical of the HR. In contrast, chloroplast ROS build‐up and LCD were significantly reduced in Xcv ‐inoculated plants expressing plastid‐targeted flavodoxin. Metabolic routes normally inhibited by pathogens were protected in the transformants, whereas other aspects of the HR, including the induction of defence‐associated genes and synthesis of salicylic and jasmonic acid, proceeded as in inoculated wild‐type plants. Therefore, ROS generated in chloroplasts during this non‐host interaction are essential for the progress of LCD, but do not contribute to the induction of pathogenesis‐related genes or other signalling components of the response.
Publikation
Ziegler, J.; Brandt, W.; Geißler, R.; Facchini, P. J.;Removal of Substrate Inhibition and Increase in Maximal Velocity in the Short Chain Dehydrogenase/Reductase Salutaridine Reductase Involved in Morphine BiosynthesisJ. Biol. Chem.28426758-26767(2009)DOI: 10.1074/jbc.M109.030957
Salutaridine reductase (SalR, EC 1.1.1.248) catalyzes the stereospecific reduction of salutaridine to 7(S)-salutaridinol in the biosynthesis of morphine. It belongs to a new, plant-specific class of short-chain dehydrogenases, which are characterized by their monomeric nature and increased length compared with related enzymes. Homology modeling and substrate docking suggested that additional amino acids form a novel α-helical element, which is involved in substrate binding. Site-directed mutagenesis and subsequent studies on enzyme kinetics revealed the importance of three residues in this element for substrate binding. Further replacement of eight additional residues led to the characterization of the entire substrate binding pocket. In addition, a specific role in salutaridine binding by either hydrogen bond formation or hydrophobic interactions was assigned to each amino acid. Substrate docking also revealed an alternative mode for salutaridine binding, which could explain the strong substrate inhibition of SalR. An alternate arrangement of salutaridine in the enzyme was corroborated by the effect of various amino acid substitutions on substrate inhibition. In most cases, the complete removal of substrate inhibition was accompanied by a substantial loss in enzyme activity. However, some mutations greatly reduced substrate inhibition while maintaining or even increasing the maximal velocity. Based on these results, a double mutant of SalR was created that exhibited the complete absence of substrate inhibition and higher activity compared with wild-type SalR.
Publikation
Ziegler, J.; Facchini, P. J.; Geißler, R.; Schmidt, J.; Ammer, C.; Kramell, R.; Voigtländer, S.; Gesell, A.; Pienkny, S.; Brandt, W.;Evolution of morphine biosynthesis in opium poppyPhytochemistry701696-1707(2009)DOI: 10.1016/j.phytochem.2009.07.006
Benzylisoquinoline alkaloids (BIAs) are a group of nitrogen-containing plant secondary metabolites comprised of an estimated 2500 identified structures. In BIA metabolism, (S)-reticuline is a key branch-point intermediate that can be directed into several alkaloid subtypes with different structural skeleton configurations. The morphinan alkaloids are one subclass of BIAs produced in only a few plant species, most notably and abundantly in the opium poppy (Papaver somniferum). Comparative transcriptome analysis of opium poppy and several other Papaver species that do not accumulate morphinan alkaloids showed that known genes encoding BIA biosynthetic enzymes are expressed at higher levels in P. somniferum. Three unknown cDNAs that are co-ordinately expressed with several BIA biosynthetic genes were identified as enzymes in the pathway. One of these enzymes, salutaridine reductase (SalR), which is specific for the production of morphinan alkaloids, was isolated and heterologously overexpressed in its active form not only from P. somniferum, but also from Papaver species that do not produce morphinan alkaloids. SalR is a member of a class of short chain dehydrogenase/reductases (SDRs) that are active as monomers and possess an extended amino acid sequence compared with classical SDRs. Homology modelling and substrate docking revealed the substrate binding site for SalR. The amino acids residues conferring salutaridine binding were compared to several members of the SDR family from different plant species, which non-specifically reduce (−)-menthone to (+)-neomenthol. Previously, it was shown that some of these proteins are involved in plant defence. The recruitment of specific monomeric SDRs from monomeric SDRs involved in plant defence is discussed.
Publikation
Widjaja, I.; Naumann, K.; Roth, U.; Wolf, N.; Mackey, D.; Dangl, J. L.; Scheel, D.; Lee, J.;Combining subproteome enrichment and Rubisco depletion enables identification of low abundance proteins differentially regulated during plant defenseProteomics9138-147(2009)DOI: 10.1002/pmic.200800293
Transgenic Arabidopsis conditionally expressing the bacterial avrRpm1 type III effector under the control of a dexamethasone‐responsive promoter were used for proteomics studies. This model system permits study of an individual effector without interference from additional bacterial components. Coupling of different prefractionation approaches to high resolution 2‐DE facilitated the discovery of low abundance proteins – enabling the identification of proteins that have escaped detection in similar experiments. A total of 34 differentially regulated protein spots were identified. Four of these (a remorin, a protein phosphatase 2C (PP2C), an RNA‐binding protein, and a C2‐domain‐containing protein) are potentially early signaling components in the interaction between AvrRpm1 and the cognate disease resistance gene product, resistance to Pseudomonas syringae pv. maculicola 1 (RPM1). For the remorin and RNA‐binding protein, involvement of PTM and post‐transcriptional regulation are implicated, respectively.