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Extremely Thermoacidophilic Metallosphaera Species Mediate Mobilization and Oxidation of Vanadium and Molybdenum Oxides.

ABSTRACT: Certain species from the extremely thermoacidophilic genus Metallosphaera directly oxidize Fe(II) to Fe(III), which in turn catalyzes abiotic solubilization of copper from chalcopyrite to facilitate recovery of this valuable metal. In this process, the redox status of copper does not change as it is mobilized. Metallosphaera species can also catalyze the release of metals from ores with a change in the metal's redox state. For example, Metallosphae ra sedula catalyzes the mobilization of uranium from the solid oxide U3O8, concomitant with the generation of soluble U(VI). Here, the mobilization of metals from solid oxides (V2O3, Cu2O, FeO, MnO, CoO, SnO, MoO2, Cr2O3, Ti2O3, and Rh2O3) was examined for M. sedula and M. prunae at 70°C and pH 2.0. Of these oxides, only V and Mo were solubilized, a process accelerated in the presence of FeCl3 However, it was not clear whether the solubilization and oxidation of these metals could be attributed entirely to an Fe-mediated indirect mechanism. Transcriptomic analysis for growth on molybdenum and vanadium oxides revealed transcriptional patterns not previously observed for growth on other energetic substrates (i.e., iron, chalcopyrite, organic compounds, reduced sulfur compounds, and molecular hydrogen). Of particular interest was the upregulation of Msed_1191, which encodes a Rieske cytochrome b 6 fusion protein (Rcbf, referred to here as V/MoxA) that was not transcriptomically responsive during iron biooxidation. These results suggest that direct oxidation of V and Mo occurs, in addition to Fe-mediated oxidation, such that both direct and indirect mechanisms are involved in the mobilization of redox-active metals by Metallosphaera species.IMPORTANCE In order to effectively leverage extremely thermoacidophilic archaea for the microbially based solubilization of solid-phase metal substrates (e.g., sulfides and oxides), understanding the mechanisms by which these archaea solubilize metals is important. Physiological analysis of Metallosphaera species growth in the presence of molybdenum and vanadium oxides revealed an indirect mode of metal mobilization, catalyzed by iron cycling. However, since the mobilized metals exist in more than one oxidation state, they could potentially serve directly as energetic substrates. Transcriptomic response to molybdenum and vanadium oxides provided evidence for new biomolecules participating in direct metal biooxidation. The findings expand the knowledge on the physiological versatility of these extremely thermoacidophilic archaea.

SUBMITTER: Wheaton GH

PROVIDER: S-EPMC6384102 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

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Extremely Thermoacidophilic Metallosphaera Species Mediate Mobilization and Oxidation of Vanadium and Molybdenum Oxides.

Wheaton Garrett H GH Vitko Nicholas P NP Counts James A JA Dulkis Jessica A JA Podolsky Igor I Mukherjee Arpan A Kelly Robert M RM

Applied and environmental microbiology 20190220 5

Certain species from the extremely thermoacidophilic genus Metallosphaera directly oxidize Fe(II) to Fe(III), which in turn catalyzes abiotic solubilization of copper from chalcopyrite to facilitate recovery of this valuable metal. In this process, the redox status of copper does not change as it is mobilized. Metallosphaera species can also catalyze the release of metals from ores with a change in the metal's redox state. For example, Metallosphaera sedula catalyzes ...[more]

PMID: 30578261

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Project description:Thermoacidophilic archaea are found in heavy metal-rich environments and, in some cases, these microorganisms are causative agents of metal mobilization through cellular processes related to their bioenergetics. Given the nature of their habitats, these microorganisms must deal with the potentially toxic effect of heavy metals. Here, we show that two thermoacidophilic Metallosphaera species with nearly identical (99.99%) genomes differed significantly in their sensitivity and reactivity to uranium. M. prunae, isolated from a smoldering heap on a uranium mine in Thuringen, Germany, could be viewed as a M-bM-^@M-^\spontaneous mutantM-bM-^@M-^] of M. sedula, an isolate from Pisciarelli solfatara near Naples, Italy. M. prunae tolerated U3O8 and U(VI) to a much greater extent than M. sedula. Within 15 minutes following exposure to M-bM-^@M-^\U(VI) shockM-bM-^@M-^], M. sedula, and not M. prunae, exhibited transcriptomic features associated with severe stress response. Furthermore, within 15 minutes post-U(VI) shock, M. prunae, and not M. sedula, showed evidence of substantial degradation of cellular RNA. This suggested that transcriptional and translational processes were aborted as a dynamic mechanism for resisting U toxicity; by 60 minutes post-U(VI) shock, RNA integrity in M. prunae recovered, and known modes for heavy metal resistance were activated. In addition, M. sedula rapidly oxidized solid U3O8 to soluble U(VI) for bioenergetic purposes, a chemolithoautotrophic feature not previously reported. M. prunae, however, did not solubilize solid U3O8 to any significant extent, thereby not exacerbating U(VI) toxicity. These results point to uranium extremophily as an adaptive, rather than intrinsic, feature for Metallosphaera species, driven by environmental factors. The study comprises 9 Samples, described in detail below. MprAU_MseAU: Transcriptional analysis of the response of Metallosphaera prunae (Mpr) and Metallosphaera sedula(Mse) to chemolithoautotrophic conditions (0.1 wt% Uranium octaoxide with CO2 supplementation in headspace). This experiment was done to identify the key terminal oxidases which responded to a Uranium oxide while doing inter-species comparison between Mpr and Mse. Transcriptional response of the terminal oxidase clusters proved that certain key genes play a role in the vastly different physiologies of these two species. MprN_MprU60: Transcriptional analysis of the response of Metallosphaera prunae (Mpr) to 60 min of Uranium shock. This experiment was done to analyze the differential transcription of Mpr cells challenged with 1 mM uranyl acetate shock (U shock) compared to normal growth. The Uranium cultures were harvested 60 min after the shock. MprN_MseN: Differential transcription of Metallosphaera species under normal growth conditions. This experiment was done to analyze the differential transcription of Mpr cells compared with Mse cells at mid log phase. MprN_MprU3h: Transcriptional response of Metallosphaera prunae (Mpr) to 3h of Uranium shock compared to normal growth. This experiment was done to analyze the differential transcription of Mpr cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 3 h after the shock. MseN_MseU15: Transcriptional response of Metallosphaera sedula (Mse) to 15 min of Uranium shock. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) compared to normal growth. The Uranium cultures were harvested 15 min after the shock. MseN_MseU60: Transcriptional response of Metallosphaera sedula to 60 min of Uranium shock. Mse cells were grown upto mid log phase after which the cells were subjected to U shock and harvested 60 min later. Biological repeats were done for both experimental conditions. MseN_MseU3h: Transcriptional response of Metallosphaera sedula (Mse) to 3h of Uranium shock compared to normal growth. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 3 h after the shock. MseU15_MseU60: Transcriptional response of Metallosphaera sedula to 15 min of Uranium shock compared with 60 min of Uranium shock. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 15 min and 60 min after the shock. MprU3h_MseU3h: Differential transcription of Metallosphaera cells under Uranium shock. This experiment was done to analyze the differential transcription of Metallosphaera sedula (Mse) and Metallosphaera prunae (Mpr) challenged with 1 mM uranyl acetate.

Project description:UnlabelledThe extremely thermoacidophilic archaeon Metallosphaera sedula mobilizes metals by novel membrane-associated oxidase clusters and, consequently, requires metal resistance strategies. This issue was examined by "shocking" M. sedula with representative metals (Co(2+), Cu(2+), Ni(2+), UO2 (2+), Zn(2+)) at inhibitory and subinhibitory levels. Collectively, one-quarter of the genome (554 open reading frames [ORFs]) responded to inhibitory levels, and two-thirds (354) of the ORFs were responsive to a single metal. Cu(2+) (259 ORFs, 106 Cu(2+)-specific ORFs) and Zn(2+) (262 ORFs, 131 Zn(2+)-specific ORFs) triggered the largest responses, followed by UO2 (2+) (187 ORFs, 91 UO2 (2+)-specific ORFs), Ni(2+) (93 ORFs, 25 Ni(2+)-specific ORFs), and Co(2+) (61 ORFs, 1 Co(2+)-specific ORF). While one-third of the metal-responsive ORFs are annotated as encoding hypothetical proteins, metal challenge also impacted ORFs responsible for identifiable processes related to the cell cycle, DNA repair, and oxidative stress. Surprisingly, there were only 30 ORFs that responded to at least four metals, and 10 of these responded to all five metals. This core transcriptome indicated induction of Fe-S cluster assembly (Msed_1656-Msed_1657), tungsten/molybdenum transport (Msed_1780-Msed_1781), and decreased central metabolism. Not surprisingly, a metal-translocating P-type ATPase (Msed_0490) associated with a copper resistance system (Cop) was upregulated in response to Cu(2+) (6-fold) but also in response to UO2 (2+) (4-fold) and Zn(2+) (9-fold). Cu(2+) challenge uniquely induced assimilatory sulfur metabolism for cysteine biosynthesis, suggesting a role for this amino acid in Cu(2+) resistance or issues in sulfur metabolism. The results indicate that M. sedula employs a range of physiological and biochemical responses to metal challenge, many of which are specific to a single metal and involve proteins with yet unassigned or definitive functions.ImportanceThe mechanisms by which extremely thermoacidophilic archaea resist and are negatively impacted by metals encountered in their natural environments are important to understand so that technologies such as bioleaching, which leverage microbially based conversion of insoluble metal sulfides to soluble species, can be improved. Transcriptomic analysis of the cellular response to metal challenge provided both global and specific insights into how these novel microorganisms negotiate metal toxicity in natural and technological settings. As genetics tools are further developed and implemented for extreme thermoacidophiles, information about metal toxicity and resistance can be leveraged to create metabolically engineered strains with improved bioleaching characteristics.

Project description:The exploration of novel hosts with the ability to assimilate formic acid, a C1 substrate that can be produced from renewable electrons and CO2, is of great relevance for developing novel and sustainable biomanufacturing platforms. Formatotrophs can use formic acid or formate as a carbon and/or reducing power source. Formatotrophy has typically been studied in neutrophilic microorganisms because formic acid toxicity increases in acidic environments below the pKa of 3.75 (25°C). Because of this toxicity challenge, utilization of formic acid as either a carbon or energy source has been largely unexplored in thermoacidophiles, species that possess the ability to produce a variety of metabolites and enzymes of high biotechnological relevance. Here we investigate the capacity of several thermoacidophilic archaea species from the Sulfolobales order to tolerate and metabolize formic acid. Metallosphaera prunae, Sulfolobus metallicus and Sulfolobus acidocaldarium were found to metabolize and grow with 1-2 mM of formic acid in batch cultivations. Formic acid was co-utilized by this species alongside physiological electron donors, including ferrous iron. To enhance formic acid utilization while maintaining aqueous concentrations below the toxicity threshold, we developed a bioreactor culturing method based on a sequential formic acid feeding strategy. By dosing small amounts of formic acid sequentially and feeding H2 as co-substrate, M. prunae could utilize a total of 16.3 mM of formic acid and grow to higher cell densities than when H2 was supplied as a sole electron donor. These results demonstrate the viability of culturing thermoacidophilic species with formic acid as an auxiliary substrate in bioreactors to obtain higher cell densities than those yielded by conventional autotrophic conditions. Our work underscores the significance of formic acid metabolism in extreme habitats and holds promise for biotechnological applications in the realm of sustainable energy production and environmental remediation.

Dataset Information

Extremely Thermoacidophilic Metallosphaera Species Mediate Mobilization and Oxidation of Vanadium and Molybdenum Oxides.

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Extremely Thermoacidophilic <i>Metallosphaera</i> Species Mediate Mobilization and Oxidation of Vanadium and Molybdenum Oxides.

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