ABSTRACT: Desulfitobacterium hafniense DCB-2 Gene Expression Differences from Pyruvate Fermentation when Grown with Ferric Citrate, Selenate, Nitrate, or Uranyl Acetate
Project description:Desulfitobacterium hafniense DCB-2, a halo-respirer and versatile inorganic ion reducer, grows in hazardous waste contaminated environments. M-BM- Gene expression changes were determined when the strain was grown in the presence of high concentrations of Fe(+3), Se(+6), U(+6), or NO3(-) as alternative electron acceptors. (Thesis by Christina L. Harzman at Michigan State University, 2009) RNA was used to directly compare gene expression in control samples of pyruvate fermentation (20 mM) to fermentation in the presence inorganic electron acceptors or respiration-inducing conditions. M-BM- The inorganic electron acceptors added to media for fermentation were 1 mM selenate or 0.5 mM uranyl acetate. The respiration-inducing conditions were 50 mM ferric citrate (20 mM lactate carbon source) or 10 mM nitrate (20 mM lactate carbon source). Microarray hybridizations were performed with a dye swap and 3 biological replicates for a total of 6 slides per condition for comparison.
Project description:Desulfitobacterium hafniense was originally isolated for its ability to use organohalogens as terminal electron acceptors in the process of organohalide respiration (OHR). However, in contrast to obligate OHR bacteria, Desulfitobacterium spp. show a highly versatile energy metabolism with the capacity to use many possible electron donors and acceptors and to grow fermentatively also. The sequencing of several Desulfitobacterium genomes displaying numerous and apparently redundant members of redox enzyme families has confirmed their large metabolic potential. Nonetheless, the enzymes responsible for many metabolic traits are not yet identified. In the present work, we conducted an extended proteomic study by comparing the proteomes of D. hafniense strain DCB-2 cultivated in different combinations of electron donors and acceptors, triggering five alternative respiratory metabolisms that include OHR, as well as fermentation. The Tandem Mass Tag labelling proteomic approach allowed us to identify almost 60% of the predicted proteome of strain DCB-2 in all six growth conditions and to quantify the relative abundance of 2796 proteins. This dataset was analysed in order to highlight the proteins that were up-regulated in one or a subset of growth conditions to identify possible key players in the different energy metabolisms. The addition of sodium sulfide as reducing agent in the medium – a very widespread practice in the cultivation of strictly anaerobic bacteria – triggered the expression of the dissimilatory sulfite reduction pathway in relatively less favourable conditions such as fermentative growth on pyruvate, respiration with H2 as electron donor and OHR conditions. The presence of H2, CO2 and acetate in the medium induced several metabolic pathways involved in carbon metabolism including the Wood-Ljungdahl pathway and two pathways related to the fermentation of butyrate that rely on electron-bifurcating enzymes. While the predicted fumarate reductase appears to be constitutively expressed, a new lactate dehydrogenase and lactate transporters were identified. Finally, the OHR metabolism with 3-chloro-4-hydroxyphenylacetate as electron acceptor strongly induced proteins encoded in several reductive dehalogenase gene clusters including four new proteins, the function of which is discussed. Overall, we believe that this extended proteomic database represents a new landmark in understanding the metabolic versatility of Desulfitobacterium spp. and provides a solid basis for addressing present and future research questions.
Project description:Desulfitobacterium hafniense DCB-2, a halo-respirer and versatile inorganic ion reducer, grows in hazardous waste contaminated environments. Gene expression changes were determined when the strain was grown in the presence of high concentrations of Fe(+3), Se(+6), U(+6), or NO3(-) as alternative electron acceptors. (Thesis by Christina L. Harzman at Michigan State University, 2009)
Project description:The physiology of D. hafniense strain Y51 under close to natural conditions were examined: D. hafniense Y51 was subjected to a variety of limiting conditions (carbon or electron acceptor) or ammonium deprivation, in chemostat. Its physiology was assessed by transcriptomic and proteomic analyses and compared to that in batch culture. Electron donor/acceptor-limited growth conditions cause a switch to alternative metabolic pathways allowing the organism with a capacity to survive in polluted environments and with a readiness to use alternative electron acceptors such as PCE; (ii) nitrogen limitation induces nitrogen fixation implying a robustness vis-a-vis ammonia shortage; (iii) limiting conditions trigger a general stress response that should enable the organism to cope with even more environmental constraints.
