84 research outputs found
On the diverse bonding situations in nanostructures : an ab initio computational study
This computational study investigates diverse bonding situations in nanostructures (carbon nanotubes, fullerenes, metal compounds) spanning a broad range of energies. Weak, dispersive interactions and covalent metal-ligand and metal-metal bonding are examined. The results of efficient density functional calculations are compared to those of correlated wavefunction calculations on model systems. This rigorous validation is crucial in evaluating the balance between computational cost and accuracy
Development and validation of a microsatellite marker-based method for tracing infections by Microsporum canis.
The Structural Basis of Peptide Binding at Class A G Protein-Coupled Receptors
G protein-coupled receptors (GPCRs) represent the largest membrane protein family and a significant target class for therapeutics. Receptors from GPCRs’ largest class, class A, influence virtually every aspect of human physiology. About 45% of the members of this family endogenously bind flexible peptides or peptides segments within larger protein ligands. While many of these peptides have been structurally characterized in their solution state, the few studies of peptides in their receptor-bound state suggest that these peptides interact with a shared set of residues and undergo significant conformational changes. For the purpose of understanding binding dynamics and the development of peptidomimetic drug compounds, further studies should investigate the peptide ligands that are complexed to their cognate receptor
Characterization of a membrane-bound C-glucosyltransferase responsible for carminic acid biosynthesis in Dactylopius coccus Costa
Carminic acid is a widely applied red colorant that is still harvested from insects because its biosynthesis is not fully understood. Here, the authors identify and characterize a membrane-bound C-glucosyltransferase catalyzing the final step during carminic acid biosynthesis
The Antibacterial Protein Lysozyme Identified as the Termite Egg Recognition Pheromone
Social insects rely heavily on pheromone communication to maintain their sociality. Egg protection is one of the most fundamental social behaviours in social insects. The recent discovery of the termite-egg mimicking fungus ‘termite-ball’ and subsequent studies on termite egg protection behaviour have shown that termites can be manipulated by using the termite egg recognition pheromone (TERP), which strongly evokes the egg-carrying and -grooming behaviours of workers. Despite the great scientific and economic importance, TERP has not been identified because of practical difficulties. Herein we identified the antibacterial protein lysozyme as the TERP. We isolated the target protein using ion-exchange and hydrophobic interaction chromatography, and the MALDI-TOF MS analysis showed a molecular size of 14.5 kDa. We found that the TERP provided antibacterial activity against a gram-positive bacterium. Among the currently known antimicrobial proteins, the molecular size of 14.5 kDa limits the target to lysozyme. Termite lysozymes obtained from eggs and salivary glands, and even hen egg lysozyme, showed a strong termite egg recognition activity. Besides eggs themselves, workers also supply lysozyme to eggs through frequent egg-grooming, by which egg surfaces are coated with saliva containing lysozyme. Reverse transcript PCR analysis showed that mRNA of termite lysozyme was expressed in both salivary glands and eggs. Western blot analysis confirmed that lysozyme production begins in immature eggs in queen ovaries. This is the first identification of proteinaceous pheromone in social insects. Researchers have focused almost exclusively on hydrocarbons when searching for recognition pheromones in social insects. The present finding of a proteinaceous pheromone represents a major step forward in, and result in the broadening of, the search for recognition pheromones. This novel function of lysozyme as a termite pheromone illuminates the profound influence of pathogenic microbes on the evolution of social behaviour in termites
Chemical warfare between leafcutter ant symbionts and a co-evolved pathogen
Acromyrmex leafcutter ants form a mutually beneficial symbiosis with the fungus Leucoagaricus gongylophorus and with Pseudonocardia bacteria. Both are vertically transmitted and actively maintained by the ants. The fungus garden is manured with freshly cut leaves and provides the sole food for the ant larvae, while Pseudonocardia cultures are reared on the ant-cuticle and make antifungal metabolites to help protect the cultivar against disease. If left unchecked, specialized parasitic Escovopsis fungi can overrun the fungus-garden and lead to colony collapse. We report that Escovopsis upregulates the production of two specialized metabolites when it infects the cultivar. These compounds inhibit Pseudonocardia and one, shearinine D, also reduces worker behavioral defences and is ultimately lethal when it accumulates in ant tissues. Our results are consistent with an active evolutionary arms race between Pseudonocardia and Escovopsis, which modifies both bacterial and behavioral defences such that colony collapse is unavoidable once Escovopsis infections escalate
Molekulare und Chemische Analysen zur Biogenese
0\. Title Page and Table of Contents
1\. General Introduction and Thesis Outline 1
2\. Anthraquinones as Defensive Compounds in Eggs of Galerucini Leaf Beetles:
Biosynthesis by the beetle? 17
3\. Presence of Wolbachia in Insect Eggs Containing Antimicrobially Active
Anthraquinones 33
4\. Different Polyketide Folding Modes Converge to an Identical Molecular
Architecture 49
5\. Defensive Compounds in Insect Eggs: Are Anthraquinones Produced during
Egg Development 61
6\. Search for Genes Involved in Anthraquinone Biosynthesis in Galeruca
tanaceti 69
7\. Polyketides in Insects 78
8\. Summary 121
9\. Zusammenfassung 125
DanksagungAnthraquinones and anthrones that are not sequestered from food are unusual
compounds in insects and only found in leaf beetles of the tribe Galerucini
and in scale insects. The major host plants of Galerucini do not contain
anthraquinones. Thus, these polyketides might be either produced by the beetle
itself or by endosymbiotic microorganisms.
The major goal of this thesis was to elucidate the origin of anthrones and
anthraquinones in Galerucini leaf beetles with molecular and chemical methods.
The tansy leaf beetle, Galeruca tanaceti, was taken as a model Galerucini
species containing chrysophanol and chrysazin and the anthrones chrysarobin
and dithranol.
Endosymbiotic microorganisms were searched in eggs of G. tanaceti. No
endosymbiotic bacteria were found except of Wolbachia. This alpha-
proteobacterium was absent in the elm leaf beetle (Xanthogaleruca luteola), a
closely related Galerucini species containing anthrones/ anthraquinones.
However, Wolbachia were present in the anthraquinone-free alder leaf beetle
(Agelastica alni). Neither could fungi responsible for anthraquinone
production be detected in eggs of G. tanaceti. Furthermore, treatment of adult
beetles with antibiotics did not block anthraquinone biosynthesis. This
finding supported the hypothesis that no endosymbiotic microorganisms are
responsible for anthraquinone production in Galerucini.
To elucidate the origin of anthraquinones in G. tanaceti the folding mode of
the polyketide chain (octaketide) leading to chrysophanol was measured with
NMR techniques. Prokaryotic organisms have another folding mode (type S) than
eukaryotic ones (type F). Chrysophanol present in larvae was shown to be
synthesised via the eukaryotic folding mode F. Thus, only the beetle itself or
a fungus can act as anthraquinone producer. Since no endosymbiotic fungi were
detected with molecular techniques, it is considered most likely that the
beetle itself is able to produce anthrones and anthraquinones.
Knowledge of the anthraquinone production site in G. tanaceti might help to
find a prove who (beetle or endosymbiont) produces the anthraquinones. The
quantities of some anthrones and anthraquinones decreased significantly during
egg development, while others stayed unchanged. These results clearly showed
that anthrones and anthraquinones are rather metabolised than produced by the
embryo within the eggs. Therefore, the polyketides are transferred by the
mother into the eggs and are produced within the female beetle.
In a final approach, we searched for genes encoding polyketide synthases
(PKS), i.e., enzymes catalysing biosynthesis of polyketides like
anthraquinones. No PKS gene responsible for the biogenesis of
1,8-dihydroxylated anthraquinones could be detected so far.
Numerous other defensive components and pheromones of insects have been
suggested to be polyketides. With the exception of PKS for the polyketide
pederin produced by endosymbiotic bacteria, no enzymes responsible for
polyketide biosynthesis have been isolated from insects. The precursors of
components produced via the polyketide pathway are very similar to those
produced via the fatty acid pathway. A brief overview of insect defensive and
pheromonal components with (putative) polyketide origin is given. Furthermore,
similarities and differences of PKS and fatty acid synthases (FAS) are
highlighted.Anthrachinone und Anthrone, die nicht aus der Nahrung sequestriert werden,
sind ungewöhnliche Substanzen in Insekten und wurden bisher nur in Blattkäfern
der Tribus Galerucini und in Schildläusen gefunden. Die Hauptfutterpflanzen
der Galerucini enthalten keine Anthrachinone. Daher werden diese Polyketide
entweder vom Käfer selbst oder von endosymbiotischen Mikroorganismen
produziert.
Das Hauptthema dieser Arbeit beschäftigte sich mit der Untersuchung des
Ursprungs der Anthrone und Anthrachinone in Blattkäfern der Tribus Galerucini.
Dafür wurden molekulare und chemische Methoden verwendet. Als Modellart eines
Blattkäfers der Galerucini wurde der Rainfarnblattkäfer (Galeruca tanaceti)
verwendet, der Chrysophanol und Chrysazin und die Anthrone Chrysarobin und
Dithranol enthält.
In den Eiern von G. tanaceti wurde nach endosymbiotischen Mikroorganismen
gesucht. Es konnten keine endosymbiotischen Bakterien außer Wolbachia gefunden
werden. Dieses alpha-Proteobakterium konnte im Ulmenblattkäfer (Xanthogaleruca
luteola), einer zu den Galerucini nahe verwandten Art, nicht nachgewiesen
werden. Im Gegensatz dazu wurden Wolbachien im anthrachinon-freien
Erlenblattkäfer (Agelastica alni) gefunden. Außerdem konnten in den Eiern von
G. tanaceti keine Pilze, die für eine Anthrachinonproduktion in Frage kämen,
nachgewiesen werden. Zusätzlich konnte gezeigt werden, dass eine Behandlung
von adulten Käfern mit Antibiotika die Anthrachinonsynthese nicht blockiert.
Diese Ergebnisse stützen die Hypothese, dass keine endosymbiotischen
Mikroorganismen für die Anthrachinonproduktion in den Galerucini
verantwortlich sind.
Um den Ursprung der Anthrachinone in G. tanaceti aufzudecken wurde der
Faltungstyp der Polyketidkette (Oktaketid), die zum Chrysophanol führt, mit
NMR Methoden untersucht. Prokaryoten haben einen anderen Faltungstyp (Typ S)
als Eukaryoten (Type F). Es konnte gezeigt werden, dass Chrysophanol aus
Larven über den eukaryotischen Faltungstyp F synthetisiert wird. Somit kommt
nur der Käfer selbst oder ein Pilz als Anthrachinonproduzent in Frage. Da mit
den molekularen Methoden keine Pilze gefunden wurden, ist eine Produktion der
Anthrone und Anthrachinone vom Käfer selbst sehr wahrscheinlich.
Kenntnisse über den Produktionsort der Anthrachinone in G. tanaceti könnten
bei der Suche nach einer Bestätigung, wer diese produziert (Käfer oder
Endosymbiont), hilfreich sein. Einige Anthron- und Anthrachinonmengen nahmen
signifikant während der Eientwicklung ab, während andere Mengen unverändert
blieben. Diese Ergebnisse zeigten, dass die Anthrone und Anthrachinone eher
vom Embryo im Ei metabolisiert als von ihm produziert wurden. Die im
weiblichen Käfer produzierten Polyketide wurden daher in die Eier
transferiert.
Abschließend wurde nach Genen für eine Polyketidsynthase (PKS) gesucht. PKS
katalysieren die Biosynthese von Polyketiden wie z.B. der Anthrachinone. Es
konnten bisher keine PKS Gene gefunden werden, die für eine Biogenese der
1,8-dihydroxylierten Anthrachinone verantwortlich sein könnten.
Bei Insekten sind zahlreiche defensive Substanzen und Pheromone bekannt, die
möglicherweise ebenfalls Polyketide sind. Bis auf die PKS für das Polyketid
Pederin produziert von Endosymbionten wurde noch kein Enzym für die
Polyketidsynthese aus Insekten isoliert. Die Bausteine von Substanzen, die
über den Polyketidweg produziert werden, sind den Bausteinen von Substanzen
sehr ähnlich, die über den Fettsäureweg produziert werden. Zuerst werden die
defensiven Substanzen und die Pheromone mit möglichem Polyketidursprung kurz
zusammengefasst. Im weiteren werden die Gemeinsamkeiten und Unterschiede von
PKS und Fettsäuresynthasen (FAS) beschrieben
The Structural Basis of Peptide Binding at Class A G Protein-Coupled Receptors
G protein-coupled receptors (GPCRs) represent the largest membrane protein family and a significant target class for therapeutics. Receptors from GPCRs’ largest class, class A, influence virtually every aspect of human physiology. About 45% of the members of this family endogenously bind flexible peptides or peptides segments within larger protein ligands. While many of these peptides have been structurally characterized in their solution state, the few studies of peptides in their receptor-bound state suggest that these peptides interact with a shared set of residues and undergo significant conformational changes. For the purpose of understanding binding dynamics and the development of peptidomimetic drug compounds, further studies should investigate the peptide ligands that are complexed to their cognate receptor
The Structural Basis of Peptide Binding at Class A G Protein-Coupled Receptors
G protein-coupled receptors (GPCRs) represent the largest membrane protein family and a significant target class for therapeutics. Receptors from GPCRs’ largest class, class A, influence virtually every aspect of human physiology. About 45% of the members of this family endogenously bind flexible peptides or peptides segments within larger protein ligands. While many of these peptides have been structurally characterized in their solution state, the few studies of peptides in their receptor-bound state suggest that these peptides interact with a shared set of residues and undergo significant conformational changes. For the purpose of understanding binding dynamics and the development of peptidomimetic drug compounds, further studies should investigate the peptide ligands that are complexed to their cognate receptor
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