The pentose metabolism of Archaea is largely unknown. Here, we have employed an integrated genomics approach including DNA microarray and proteomics analyses to elucidate the catabolic pathway for D-arabinose in Sulfolobus solfataricus. During growth on this sugar, a small set of genes appeared to be differentially expressed compared with growth on D-glucose. These genes were heterologously overexpressed in Escherichia coli, and the recombinant proteins were purified and biochemically studied. This showed that D-arabinose is oxidized to 2-oxoglutarate by the consecutive action of a number of previously uncharacterized enzymes, including a D-arabinose dehydrogenase, a D-arabinonate dehydratase, a novel 2-keto-3-deoxy-D-arabinonate dehydratase, and a 2,5-dioxopentanoate dehydrogenase. Promoter analysis of these genes revealed a palindromic sequence upstream of the TATA box, which is likely to be involved in their concerted transcriptional control. Integration of the obtained biochemical data with genomic context analysis strongly suggests the occurrence of pentose oxidation pathways in both Archaea and Bacteria, and predicts the involvement of additional enzyme components. Moreover, it revealed striking genetic similarities between the catabolic pathways for pentoses, hexaric acids, and hydroxyproline degradation, which support the theory of metabolic pathway genesis by enzyme recruitment.Pentose sugars are a ubiquitous class of carbohydrates with diverse biological functions. Ribose and deoxyribose are major constituents of nucleic acids, whereas arabinose and xylose are building blocks of several plant cell wall polysaccharides. Many prokaryotes, as well as yeasts and fungi, are able to degrade these polysaccharides, and use the released five-carbon sugars as a sole carbon and energy source. At present, three main catabolic pathways have been described for pentoses. The first is present in Bacteria and uses isomerases, kinases, and epimerases to convert D-and L-arabinose (Ara) and D-xylose (Xyl) into D-xylulose 5-phosphate (Fig. 1A), which is further metabolized by the enzymes of the phosphoketolase or pentose phosphate pathway. The genes encoding the pentose-converting enzymes are often located in gene clusters in bacterial genomes, for example, the araBAD operon for L-Ara (1), the xylAB operon for D-Xyl (2), and the darK-fucPIK gene cluster for D-Ara (3). The second catabolic pathway for pentoses converts D-Xyl into D-xylulose 5-phosphate as well, but the conversions are catalyzed by reductases and dehydrogenases instead of isomerases and epimerases (Fig. 1B). This pathway is commonly found in yeasts, fungi, mammals, and plants, but also in some bacteria (4 -6). In a third pathway, pentoses such as L-Ara, D-Xyl, D-ribose, and D-Ara are metabolized non-phosphorylatively to either 2-oxoglutarate (2-OG) 4 or to pyruvate and glycolaldehyde (Fig. 1C). The conversion to 2-OG, which is a tricarboxylic acid cycle intermediate, proceeds via the subsequent action of a pentose dehydrogenase, a pentonolactonase, a pentoni...

In anoxic environments such as swamps, rice fields and sludge digestors, syntrophic microbial communities are important for decomposition of organic matter to CO2 and CH4 . The most difficult step is the fermentative degradation of short-chain fatty acids such as propionate and butyrate. Conversion of these metabolites to acetate, CO2 , formate and hydrogen is endergonic under standard conditions and occurs only if methanogens keep the concentrations of these intermediate products low. Butyrate and propionate degradation pathways include oxidation steps of comparably high redox potential, i.e. oxidation of butyryl-CoA to crotonyl-CoA and of succinate to fumarate, respectively, that require investment of energy to release the electrons as hydrogen or formate. Although investigated for several decades, the biochemistry of these reactions is still not completely understood. Genome analysis of the butyrate-oxidizing Syntrophomonas wolfei and Syntrophus aciditrophicus and of the propionate-oxidizing Syntrophobacter fumaroxidans and Pelotomaculum thermopropionicum reveals the presence of energy-transforming protein complexes. Recent studies indicated that S. wolfei uses electron-transferring flavoproteins coupled to a menaquinone loop to drive butyryl-CoA oxidation, and that S. fumaroxidans uses a periplasmic formate dehydrogenase, cytochrome b:quinone oxidoreductases, a menaquinone loop and a cytoplasmic fumarate reductase to drive energy-dependent succinate oxidation. Furthermore, we propose that homologues of the Thermotoga maritima bifurcating [FeFe]-hydrogenase are involved in NADH oxidation by S. wolfei and S. fumaroxidans to form hydrogen.

In sulfate-reducing and methanogenic environments complex biopolymers are hydrolyzed and degraded by fermentative micro-organisms that produce hydrogen, carbon dioxide and short chain fatty acids. Degradation of short chain fatty acids can be coupled to methanogenesis or to sulfate-reduction. Here we study from a genome perspective why some of these micro-organisms are able to grow in syntrophy with methanogens and others are not. Bacterial strains were selected based on genome availability and upon their ability to grow on short chain fatty acids alone or in syntrophic association with methanogens. Systematic functional domain profiling allowed us to shed light on this fundamental and ecologically important question. Extra-cytoplasmic formate dehydrogenases (InterPro domain number; IPR006443), including their maturation protein FdhE (IPR024064 and IPR006452) is a typical difference between syntrophic and non-syntrophic butyrate and propionate degraders. Furthermore, two domains with a currently unknown function seem to be associated with the ability of syntrophic growth. One is putatively involved in capsule or biofilm production (IPR019079) and a second in cell division, shape-determination or sporulation (IPR018365). The sulfate-reducing bacteria Desulfobacterium autotrophicum HRM2, Desulfomonile tiedjei and Desulfosporosinus meridiei were never tested for syntrophic growth, but all crucial domains were found in their genomes, which suggests their possible ability to grow in syntrophic association with methanogens. In addition, profiling domains involved in electron transfer mechanisms revealed the important role of the Rnf-complex and the formate transporter in syntrophy, and indicate that DUF224 may have a role in electron transfer in bacteria other than Syntrophomonas wolfei as well. This article is a part of a Special Issue entitled: 18th European Bioenergetics Conference (Biochim. Biophys. Acta, Volume 1837, Issue 7, July 2014).

Transcription of genes coding for formate dehydrogenases (fdh genes) and hydrogenases (hyd genes) in Syntrophobacter fumaroxidans and Methanospirillum hungatei was studied following growth under different conditions. Under all conditions tested, all fdh and hyd genes were transcribed. However, transcription levels of the individual genes varied depending on the substrate and growth conditions. Our results strongly suggest that in syntrophically grown S. fumaroxidans cells, the [FeFe]-hydrogenase (encoded by Sfum_844-46), FDH1 (Sfum_2703-06) and Hox (Sfum_2713-16) may confurcate electrons from NADH and ferredoxin to protons and carbon dioxide to produce hydrogen and formate, respectively. Based on bioinformatic analysis, a membrane-integrated energy-converting ,-methylene-H 4 MPT reduction are the cytoplasmic [NiFe]-hydrogenase and FDH1. 16S rRNA ratios indicate that in one of the triplicate co-cultures of S. fumaroxidans and M. hungatei, less energy was available for S. fumaroxidans. This led to enhanced transcription of genes coding for the Rnf-complex (Sfum_2694-99) and of several fdh and hyd genes. The Rnf-complex probably reoxidized NADH with ferredoxin reduction, followed by ferredoxin oxidation by the induced formate dehydrogenases and hydrogenases.

Syntrophobacter fumaroxidans strain MPOBT is the best-studied species of the genus Syntrophobacter. The species is of interest because of its anaerobic syntrophic lifestyle, its involvement in the conversion of propionate to acetate, H2 and CO2 during the overall degradation of organic matter, and its release of products that serve as substrates for other microorganisms. The strain is able to ferment fumarate in pure culture to CO2 and succinate, and is also able to grow as a sulfate reducer with propionate as an electron donor. This is the first complete genome sequence of a member of the genus Syntrophobacter and a member genus in the family Syntrophobacteraceae. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 4,990,251 bp long genome with its 4,098 protein-coding and 81 RNA genes is a part of the Microbial Genome Program (MGP) and the Genomes to Life (GTL) Program project.