München, Techn. Univ., Diss., 2001. / Computerdatei im Fernzugriff.

München, Techn. Univ., Diss., 2001. / Computerdatei im Fernzugriff.

München, Techn. University, Diss., 2001.

Univ., Diss.--Hannover, 1997.

No description available.

Universiẗat-Gesamthochsch., Diss.--Paderborn, 1996.

Cultivation of a model cyanobacterium Synechocystis PCC 6803 under low light conditions in the presence of glucose and TES buffer leads to a change of the medium color from colorless to yellow. The absorption spectrum of the excreted unknown compound indicated a possible relationship to plant chlorophyll degradation products. To confirm this speculation the compound was purified by a combination of solid phase extraction and HPLC. The mass and NMR characteristics excluded its close relationship to modified tetrapyrroles, nevertheless the precise structure could not be determined by these means due to a complicated nature of the compound and its high polarity.

This thesis focuses on the nature of heme metabolism in various eukaryotes. One of the aims was the elucidation of the origin of the unique heme biosynthesis pathway in apicomplexan parasites through a comparative study of their photosynthetic relative Chromera velia combining molecular biology, biochemistry and bioinformatics approach. Using similar strategy, I have also investigated the origin and spatial organization of tetrapyrrole biosynthesis in Euglena gracilis. Based on the phylogenetic data I described the complex evolution of heme metabolism in kinetoplastid flagellates including pathogenic trypanosomes. I revealed that one of them (Phytomonas) does not require heme for viability by the combination of various biochemical and molecular biology experiments and bioinformatic analyses.

Die für Pflanzen lebenswichtige Synthese von Tetrapyrrolen bedarf einer fein justierten Anpassung an die Umweltbedingungen und erfolgt auf transkriptioneller und post-translationaler Ebene. In den Chloroplasten hat sich die Regulation von Enzymen über ihren Redox-Status als probates Mittel zur Koordination der photosynthetischen Energiegewinnung und des Metabolismus erwiesen. Die bei der Photosynthese generierten Reduktionsäquivalente werden zum Teil über die Ferredoxin-Thioredoxin-Reduktase auf eine Vielzahl plastidärer Thioredoxine (TRX) übertragen, welche Disulfidbrücken ihrer Zielproteine reduzieren können. Unter den Enzymen der Tetrapyrrolbiosynthese (TBS) wurden bisher mehrere TRX-Interaktionspartner identifiziert, darunter die Glutamyl-tRNA-Reduktase (GluTR) und die 5-Aminolävulinsäure-Dehydratase (ALAD). In Arabidopsis Mutanten, in denen die NADPH-abhängige Thioredoxin-Reduktase (NTRC) oder f- und m-Typ-TRX fehlen, konnten verringerte Chlorophyll- und Hämgehalte beobachtet werden. Diese ließen sich auf die verringerte Stabilität verschiedener TBS-Enzyme in den Mutanten zurückführen, darunter auch die GluTR und ALAD. Die Relevanz der Cysteine (Cys, C) für die Regulation der GluTR1-Stabilität wurde in vivo über transgene Arabidopsis Cys➔Serin (Ser, S)-Substitutionslinien untersucht. Dabei erwies sich GluTR1(C464S) stärker vor dem Abbau über die kaseinolytische Protease (Clp) geschützt als das WT-Protein. Eine intermolekulare Disulfidbrücke zwischen den beiden Cys464-Resten des GluTR1-Homodimers wird daher als Abbausignal postuliert. Mit Hilfe der rekombinanten ALAD1(Cys➔Ser)-Substitutionsmutanten konnte gezeigt werden, dass nicht nur die Stabilität, sondern auch die Aktivität der ALAD1 in vitro vom Redox-Status des Enzyms abhängig ist. Die ALAD1(Cys➔Ser)-Substitutionsmutanten konnten über Enzymaktivitäts- und gel shift-Assays unter oxidierenden und reduzierenden Bedingungen zur Identifizierung der redox-sensitiven Cys beitragen. / The synthesis of tetrapyrroles, such as chlorophyll, is vital for plants and requires a finetuned regulation. The control mechanisms involved in tetrapyrrole biosynthesis (TBS) take place both on transcriptional and post-translational levels. A broadly spread post-translational regulatory mechanism in the chloroplast involves the reduction of inter- or intramolecular disulfide bonds of redox-sensitive target enzymes by thioredoxins (TRX). Thereby the coupling of photosynthetic energy production with several energy-consuming metabolic processes can be accomplished. The reduction of disulfide bonds in redox-sensitive enzymes was previously shown to lead usually to their activation. Regarding the TBS, several TRX interacting proteins have been identified, including glutamyl-tRNA-reductase (GluTR) as well as the 5-aminolevulinic acid dehydratase (ALAD). Through the detailed and combined analysis of mutants with deficient NADPH-dependent thioredoxin reductase C (NTRC), TRX-f and TRX-m, a correlation became evident between decreased chlorophyll and heme levels of the mutants and lower amounts of several TBS enzymes, including GluTR and ALAD. For GluTR1, transgenic Arabidopsis cysteine (Cys, C) ➔ serine (Ser, S) substitution lines were generated and analyzed to identify the redox-sensitive Cys residues in vivo. In these studies, GluTR1(C464S) was shown to be better protected from degradation by the caseinolytic protease (Clp) than the GluTR1 WT protein. Thus, an intermolecular disulfide bond between the Cys464 residues in the dimerization domain of the GluTR1 homodimer is postulated to serve as a degradation signal under oxidizing conditions. However, it was shown by activity- and gel shift-assays with recombinant ALAD1(Cys➔Ser) substitution mutants that not only the stability, but also the in vitro activity of ALAD1 depends on the enzyme's redox state. Redox-sensitive inter- and intramolecular disulfide bridges of ALAD1 were identified among Cys71, Cys152 and Cys251.

The Small Cab-like Proteins (SCPs) in the cyanobacterium Synechocystis sp. PCC 6803 accumulate in cells grown under different stress conditions. Genes coding for SCPs have been found in all sequenced organisms performing oxygenic photosynthesis and even in the genomes of cyanophages. Deletion of multiple scp genes in Synechocystis resulted in mutants with severely impaired growth and altered pigment content. These findings indicate the importance of SCPs in photosynthesis; however, their specific function is not well understood. SCPs share a chlorophyll-binding motif with the plant light harvesting complex, suggesting that they bind chlorophyll. Here I describe my findings, which unambiguously show that SCPs are able to bind chlorophyll in vitro. Although they affect both the stoichiometric ratio of Photosystem I to II and chlorophyll stability, they do not seem to be directly involved in non-photochemical quenching. I was able to reveal the location of the SCPs within the cyanobacterial cell: in stressed cells they attach to Photosystem II in the thylakoid membrane. Furthermore, I revealed the presence of another light-harvesting like (Lil)/SCP protein in Synechocystis sp. PCC 6803. The gene, slr1544, codifying for this newly characterised LilA protein, co-transcribes together with scpD and also appears to bind to Photosystem II during stress.