Project description:The purple sulfur bacterium Allochromatium vinosum DSM 180T is one of the best studied sulfur-oxidizing anoxygenic phototrophic bacteria and has been developed into a model organism for laboratory-based studies of oxidative sulfur metabolism. Here, we took advantage of the organismM-bM-^@M-^Ys high metabolic versatility and performed whole-genome transcriptional profiling to investigate the response of A. vinosum cells upon exposure to sulfide, thiosulfate, elemental sulfur or sulfite as compared to photoorganoheterotrophic growth on malate. Differential expression (at least twofold) of 1149 genes was observed, corresponding to 30% of the A. vinosum genome. A total of 549 genes were identified for which relative transcription increased at least twofold during growth on one of the different sulfur sources while relative transcription of 599 genes decreased. A significant number of genes that were strongly induced have documented sulfur-metabolism-related functions. Among these are the dsr genes including dsrAB for dissimilatory sulfite reductase and the sgp genes for the proteins of the sulfur globule envelope thus confirming former results. In addition we were able to identify new genes encoding proteins with appropriate subcellular localization and properties to participate in oxidative dissimilatory sulfur metabolism. Two of these were chosen for inactivation and phenotypic analyses of the respective mutant strains. This approach verified the importance of the encoded proteins for the oxidation of sulfide and thereby also documented the suitability of comparative transcriptomics for the identification of new sulfur-related genes in anoxygenic phototrophic sulfur bacteria. In this study, the relative genomic expression profiles of A. vinosum DSM 180T growing photolithoautotrophically on different reduced sulfur compounds were determined in comparison to those of cells grown photoorganoheterothrophically on malate (RCV medium) at exactly the same light intensity. The malate-containing medium was supplied with 0.815 mM sulfate in order to satisfy the sulfur-requirement for biosynthesis of sulfur-containing cell constituents. Three independent photolithoautotrophic cultures each, grown on sulfide, thiosulfate or sulfite were harvested 1 h, 2 h or 7 h, respectively, after inoculation. When elemental sulfur was the substrate, four independent cultures were harvested 3 h after inoculation.

Project description:Insights into the Metabolism of Elemental Sulfur by the Hyperthermophilic Archaeon Pyrococcus furiosus: Characterization of a Coenzyme A-Dependent NAD(P)H Sulfur Oxidoreductase The hyperthermophilic archaeon Pyrococcus furiosus, uses carbohydrates as a carbon source and produces acetate, CO2 and H2 as end products. When S° is added to a growing culture, within 10 min the rate of H2 production rapidly decreases and H2S is detected. After one hour cells contain high NADPH- and coenzyme A-dependent S° reduction activity (0.7 units/mg, 85°C) located in the cytoplasm. The enzyme responsible for this activity was purified to electrophoretic homogeneity (specific activity, 100 units/mg) and is termed NAD(P)H elemental sulfur oxidoreductase (NSR). NSR is a homodimeric flavoprotein (Mr 100 kDa) and is encoded by PF1186. This was previously assigned to an enzyme that reduces coenzyme A disulfide, which is a side-reaction of NSR. Whole genome DNA microarray and quantitative PCR analyses showed that the expression of NSR is up-regulated up to 7-fold within 10 min of S° addition. This primary response to S° also involves the up-regulation (> 16-fold) of a 13 gene cluster encoding a membrane-bound oxidoreductase (MBX). MBX is proposed replace the homologous 14 gene cluster that encodes the ferredoxin-oxidizing, H2-evolving membrane-bound hydrogenase (MBH), which is down-regulated >12-fold within 10 min of S° addition. Although an activity for MBX could not be demonstrated, it is proposed to conserve energy by oxidizing ferredoxin and reducing NADP, which is used by NSR to reduce S°. A secondary response to S° is observed 30 min after S° addition and includes the up-regulation of genes encoding proteins involved in amino acid biosynthesis and iron metabolism, as well as two so-called sulfur-induced proteins, termed SipA and SipB. This novel S°-reducing system involving NSR and MBX is so far unique to the heterotrophic Thermococcales, and is in contrast to the cytochrome- and quinone-based S°-reducing system in autotrophic archaea and bacteria. Keywords: time course, kinetic, sulfur metabolism, archaea, Pyrococcus furiosus, hyperthermophile

Project description:Responses of photosynthetic organisms to sulfur starvation include (i) increasing the capacity of the cell for transporting and/or assimilating exogenous sulfate, (ii) restructuring cellular features to conserve sulfur resources, and (iii) modulating metabolic processes and rates of cell growth and division. We used microarray analyses to obtain a genome-level view of changes in mRNA abundances in the green alga Chlamydomonas reinhardtii during sulfur starvation. The work confirms and extends upon previous findings showing that sulfur deprivation elicits changes in levels of transcripts for proteins that help scavenge sulfate and economize on the use of sulfur resources. Changes in levels of transcripts encoding members of the light-harvesting polypeptide family, such as LhcSR2, suggest restructuring of the photosynthetic apparatus during sulfur deprivation. There are also significant changes in levels of transcripts encoding enzymes involved in metabolic processes (e.g., carbon metabolism), intracellular proteolysis, and the amelioration of oxidative damage; a marked and sustained increase in mRNAs for a putative vanadium chloroperoxidase and a peroxiredoxin may help prolong survival of C. reinhardtii during sulfur deprivation. Furthermore, many of the sulfur stress-regulated transcripts (encoding polypeptides associated with sulfate uptake and assimilation, oxidative stress, and photosynthetic function) are not properly regulated in the sac1 mutant of C. reinhardtii, a strain that dies much more rapidly than parental cells during sulfur deprivation. Interestingly, sulfur stress elicits dramatic changes in levels of transcripts encoding putative chloroplast-localized chaperones in the sac1 mutant but not in the parental strain. These results suggest various strategies used by photosynthetic organisms during acclimation to nutrient-limited growth. An all pairs experiment design type is where all labeled extracts are compared to every other labeled extract. Keywords: all_pairs

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Sulfur-oxidizing prototype biomass.

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