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Item Alternative sources of molybdenum for Methanococcus maripaludis and their implication for the evolution of molybdoenzymes(Springer Science and Business Media LLC, 2024-10) Payne, Devon; Keller, Lisa M.; Larson, James; Bothner, Brian; Colman, Daniel; Boyd, Eric S.Molybdoenzymes are essential in global nitrogen, carbon, and sulfur cycling. To date, the only known bioavailable source of molybdenum (Mo) is molybdate. However, in the sulfidic and anoxic (euxinic) habitats that predominate in modern subsurface environments and that were pervasive prior to Earth’s widespread oxygenation, Mo occurs as soluble tetrathiomolybdate ion and molybdenite mineral that is not known to be bioavailable. This presents a paradox for how organisms obtain Mo to support molybdoenzymes in these environments. Here, we show that tetrathiomolybdate and molybdenite sustain the high Mo demand of a model anaerobic methanogen, Methanococcus maripaludis, grown via Mo-dependent formate dehydrogenase, formylmethanofuran dehydrogenase, and nitrogenase. Cells grown with tetrathiomolybdate and molybdenite have similar growth kinetics, Mo content, and transcript levels of proteins involved in Mo transport and cofactor biosynthesis when compared to those grown with molybdate, implying similar mechanisms of transport and cofactor biosynthesis. These results help to reconcile the paradox of how Mo is acquired in modern and ancient anaerobes and provide new insight into how molybdoenzymes could have evolved prior to Earth’s oxygenation.