Publications by Colleges and Departments (MSU - Bozeman)
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Item Methyl-reducing methanogenesis by a thermophilic culture of Korarchaeia(Springer Science and Business Media LLC, 2024-07) Krukenberg, Viola; Kohtz, Anthony J.; Jay, Zackary J.; Hatzenpichler, RolandMethanogenesis mediated by archaea is the main source of methane, a strong greenhouse gas, and thus is critical for understanding Earth’s climate dynamics. Recently, genes encoding diverse methanogenesis pathways have been discovered in metagenome-assembled genomes affiliated with several archaeal phyla1,2,3,4,5,6,7. However, all experimental studies on methanogens are at present restricted to cultured representatives of the Euryarchaeota. Here we show methanogenic growth by a member of the lineage Korarchaeia within the phylum Thermoproteota (TACK superphylum)5,6,7. Following enrichment cultivation of ‘Candidatus Methanodesulfokora washburnenis’ strain LCB3, we used measurements of metabolic activity and isotope tracer conversion to demonstrate methanol reduction to methane using hydrogen as an electron donor. Analysis of the archaeon’s circular genome and transcriptome revealed unique modifications in the energy conservation pathways linked to methanogenesis, including enzyme complexes involved in hydrogen and sulfur metabolism. The cultivation and characterization of this new group of archaea is critical for a deeper evaluation of the diversity, physiology and biochemistry of methanogens.Item Impact of mineral and non-mineral sources of iron and sulfur on the metalloproteome of Methanosarcina barkeri(American Society for Microbiology, 2024-07) Larson, James; Tokmina-Lukaszewska, Monika; Payne, Devon; Spietz, Rachel L.; Fausset, Hunter; Alam, Md Gahangir; Brekke, Brooklyn K.; Pauley, Jordan; Hasenoehrl, Ethan J.; Shepard, Eric M.; Boyd, Eric S.; Bothner, BrianMethanogens often inhabit sulfidic environments that favor the precipitation of transition metals such as iron (Fe) as metal sulfides, including mackinawite (FeS) and pyrite (FeS2). These metal sulfides have historically been considered biologically unavailable. Nonetheless, methanogens are commonly cultivated with sulfide (HS-) as a sulfur source, a condition that would be expected to favor metal precipitation and thus limit metal availability. Recent studies have shown that methanogens can access Fe and sulfur (S) from FeS and FeS2 to sustain growth. As such, medium supplied with FeS2 should lead to higher availability of transition metals when compared to medium supplied with HS-. Here, we examined how transition metal availability under sulfidic (i.e., cells provided with HS- as sole S source) versus non-sulfidic (cells provided with FeS2 as sole S source) conditions impact the metalloproteome of Methanosarcina barkeri Fusaro. To achieve this, we employed size exclusion chromatography coupled with inductively coupled plasma mass spectrometry and shotgun proteomics. Significant changes were observed in the composition and abundance of iron, cobalt, nickel, zinc, and molybdenum proteins. Among the differences were alterations in the stoichiometry and abundance of multisubunit protein complexes involved in methanogenesis and electron transport chains. Our data suggest that M. barkeri utilizes the minimal iron-sulfur cluster complex and canonical cysteine biosynthesis proteins when grown on FeS2 but uses the canonical Suf pathway in conjunction with the tRNA-Sep cysteine pathway for iron-sulfur cluster and cysteine biosynthesis under sulfidic growth conditions.Item Investigating Abiotic and Biotic Mechanisms of Pyrite Reduction(Frontiers Media SA, 2022-05) Lange Spietz, Rachel K.; Payne, Devon; Kulkarni, Gargi; Metcalf, William W.; Roden, Eric E.; Boyd, Eric S.Pyrite (FeS2) has a very low solubility and therefore has historically been considered a sink for iron (Fe) and sulfur (S) and unavailable to biology in the absence of oxygen and oxidative weathering. Anaerobic methanogens were recently shown to reduce FeS2 and assimilate Fe and S reduction products to meet nutrient demands. However, the mechanism of FeS2 mineral reduction and the forms of Fe and S assimilated by methanogens remained unclear. Thermodynamic calculations described herein indicate that H2 at aqueous concentrations as low as 10–10 M favors the reduction of FeS2, with sulfide (HS–) and pyrrhotite (Fe1–xS) as products; abiotic laboratory experiments confirmed the reduction of FeS2 with dissolved H2 concentrations greater than 1.98 × 10–4 M H2. Growth studies of Methanosarcina barkeri provided with FeS2 as the sole source of Fe and S resulted in H2 production but at concentrations too low to drive abiotic FeS2 reduction, based on abiotic laboratory experimental data. A strain of M. barkeri with deletions in all [NiFe]-hydrogenases maintained the ability to reduce FeS2 during growth, providing further evidence that extracellular electron transport (EET) to FeS2 does not involve H2 or [NiFe]-hydrogenases. Physical contact between cells and FeS2 was required for mineral reduction but was not required to obtain Fe and S from dissolution products. The addition of a synthetic electron shuttle, anthraquinone-2,6-disulfonate, allowed for biological reduction of FeS2 when physical contact between cells and FeS2 was prohibited, indicating that exogenous electron shuttles can mediate FeS2 reduction. Transcriptomics experiments revealed upregulation of several cytoplasmic oxidoreductases during growth of M. barkeri on FeS2, which may indicate involvement in provisioning low potential electrons for EET to FeS2. Collectively, the data presented herein indicate that reduction of insoluble FeS2 by M. barkeri occurred via electron transfer from the cell surface to the mineral surface resulting in the generation of soluble HS– and mineral-associated Fe1–xS. Solubilized Fe(II), but not HS–, from mineral-associated Fe1–xS reacts with aqueous HS– yielding aqueous iron sulfur clusters (FeSaq) that likely serve as the Fe and S source for methanogen growth and activity. FeSaq nucleation and subsequent precipitation on the surface of cells may result in accelerated EET to FeS2, resulting in positive feedback between cell activity and FeS2 reduction.