Reversed electron transport in syntrophic degradation of glucose, butyrate and ethanol

Abstract

The anaerobic bacteria investigated in this thesis can gain energy from metabolization of their respective substrates only by close cooperation with methanogenic archaea. By fermentation of the substrate through the fermenting bacterium hydrogen is being formed, which is used by the hydrogen-scavenging partner bacterium for the production of methane. Thus a low hydrogen partial pressure is being maintained which allows the oxidation of the thermodynamically unfavourable substrate. This exceptional case of a symbiotic relationship is defined as syntrophy. In some cases, the fermenting bacterium has to invest energy in addition to shift electrons derived from oxidation processes to the redox potential of proton reduction to hydrogen, which is called reversed electron transport.The aim of this study was the biochemical characterization of the components of the reversed electron transport in the fermenting bacteria of syntrophic cocultures growing on glucose, butyrate, or ethanol. For the glucose-utilizing bacterium Bacillus sp. BoGlc83 it was shown that, besides acetate, lactate and traces of succinate could be formed during syntrophic growth. Interspecies electron transfer occurs most likely through formate. The bacterium has all glycolytic enzymes as well as all enzymes necessary for the formation of acetate and lactate from pyruvate. Therefore, a fermentation of glucose should be possible without syntrophic partner. However, the bacterium strictly depends on the presence of a methanogenic partner organism. This phenomenon could not be explained sufficiently during this study.For syntrophic oxidation of butyrate by Syntrophomonas wolfei, a novel enzyme system has been described which catalyzes the oxidation of NADH with several different electron acceptors. By inhibition of this enzyme system with trifluoperazine in vivo, it was shown that this enzyme system is essential for butyrate oxidation and regeneration of redox carriers. More detailed characterization on the basis of sequence information from the genome of S. wolfei revealed a homology of this enzyme system with the confurcating enzyme system from Thermotoga maritima, which catalyzes the concomitant oxidation of NADH and reduced ferredoxin with protons to form hydrogen. Yet, this reaction could not be shown for S. wolfei. Instead, an enzyme reaction with quinones located in the cytoplasmic membrane was postulated. This process could be driven by proton influx into the cell.In the case of syntrophic oxidation of ethanol, enzyme activities and growth yields of two different cocultures were compared. The coculture Desulfovibrio strain KoEME1 plus Methanospirillum hungatei had significantly lower growth yields compared to the coculture Pelobacter acetylenicus plus M. hungatei. This was explained by the absence of an acetylating acetaldehyde dehydrogenase in Desulfovibrio strain KoEME1. Both Desulfovibrio strain KoEME1 and Pelobacter acetylenicus showed activities of a non-acetylating und therefore non-energy conserving acetaldehyde dehydrogenase which most likely facilitates the endergonic oxidation of ethanol by lowering the intracellular concentration of acetaldehyde.