Methanogenic organisms belong to the kingdom of archaea and are widespread in anoxic environments. The process of methanogenesis is important for the global carbon cycle because it represents the terminal step in the anaerobic breakdown of organic matter. Large amounts of CH₄ escape into the atmosphere where it acts as a greenhouse gas. The central intermediate in methanogenesis is methyl-coenzyme M, which is reductively demethylated to methane. The electrons are derived from coenzyme B and the reaction leads to the formation of a heterodisulfide from CoB-SH and CoM-SH. The reduction of the heterodisulfide is an energy-conserving step in the metabolism of methylotrophic methanogens. Two proton translocating enzyme systems, the H2:heterodisulfide oxidoreductase and the F₄₂₀H₂: heterodisulfide oxidoreductase, are involved in the membrane-bound electron transfer. We are interested in the genetic, biochemical and structural analysis of the components which participate in these reactions. In this respect we focus on the characterization of the F₄₂₀H₂ dehydrogenase which is closely related to NADH dehydrogenases (complex I of respiratory chains) and we analyse the recently discovered cofactor methanophenazine which is a unique electron carrier in the cytoplasmic membrane of Methanosarcina spec.

2. Analysis of incomplete oxidation processes as performed by Gluconobacter strains

An important group of aerobic bacteria, which is particularly characterized by its ability to oxidize organic substrates incompletely, is the one of acetic acid bacteria. Especially members of the genus Gluconobacter are famous for their rapid and incomplete oxidation of a wide range of sugars and alcohols and the nearly quantitative excretion of the oxidation products into the medium. Modern fermentation processes, such as the production of L-sorbose (ascorbate synthesis), amino-L-sorbose (miglitol synthesis), gluconic acid (sequestering agent) and dihydroxyacetone are carried out with representatives of this genus. The bacterium G. oxydans not only exhibits an extraordinarily uniqueness in its biochemistry (incomplete oxidation), but also in its growth behaviour (Y_max = 2,1 g/mol O₂) and response to culture conditions (growth in media containing up to 30 % glucose and pH value below 3). This uniqueness makes it an ideal organism for microbial process development. As mentioned above G. oxydans contains a variety of sugar-oxidizing enzymes which are membrane-bound and orientated towards the periplasm. The goal of this project is to analyze the oxidative potential by biochemical methods and to extent the substrate spectrum of the organisms by molecular biological techniques.

3. Genome analysis

Major research interests are in the area of genomics and transcriptomics of Methanosarcina mazei and Gluconobacter oxydans which have been completely sequenced recently. Cells adjust to changes in their environment in several ways, including alteration of gene expression patterns. In prokaryotes, it is well established that regulation of gene expression occurs primarily via transcriptional control through interactions between promoter and transcription regulators. Transcript abundance, a function of transcript synthesis and degradation, provides an estimate of the gene product titer. Thus, accurate measurement of mRNA levels could reveal expression alterations caused by environmental challenges. Such information is vital in furthering our understanding of cellular regulations. Therefore, we focus on a genome-wide expression profiling of these organisms by using high-density DNA arrays.. This is done by creating arrays composed of PCR products from all ORFs to monitor expression in Ms. mazei and G. oxydans at single gene resolution. The experiments shall contribute to the understanding of cellular processes that directly or indirectly affect methanogenesis and incomplete oxidation, respectively.