The metallome of Methanosarcina barkeri during electron uptake from acathode

Electromethanogenesis, the cathode-dependent reduction of CO2 to CH4 by methanogens, offers a sustainable route to methane fuel. Methanosarcina barkeri lacks surface-exposed multiheme cytochromes for extracellular electron transfer (EET). Instead, we recently showed that surface-bound G-quadruplex ribonucleic acids (G4-RNA) are required for EET, yet how electrons traverse this extracellular matrix remains unresolved. Here, we quantified metal accumulation during cathodic growth by inductively coupled plasma mass spectrometry in cells grown on cathodes poised at -430 mV versus the standard hydrogen electrode, using acetate-grown cells, open-circuit controls and abiotic cathodes for comparison. Cathode-grown M. barkeri showed CH4 buildup attributable to cathodic electrons (2.1 {+/-} 0.8% CH4), whereas open-circuit controls showed negligible increase (0.26 {+/-} 0.15% CH4). Cathode-bound cells exhibited [~]55-fold enrichment in Co, Ni and Mo, and 5- to 21-fold enrichment in Cu, Zn, and Fe relative to acetate-grown cells; neither acetate-grown cells nor abiotic cathodes accumulated metals. To resolve where metals reside, we mapped the elemental distribution in acetate-grown cells by scanning transmission electron microscopy-energy dispersive X-ray spectroscopy and high-resolution nano X-ray fluorescence. Fe co-localized with phosphorus in intracellular storage bodies, whereas Co and Zn localized within the extracellular capsule. Together, these data indicate selective metal sequestration during electromethanogenesis and raise the possibility that certain metals associate with G4-RNA and/or the methanochondroitin matrix to support charge transfer at the cell surface. This metalomic fingerprint provides a new proxy for dissecting archaeal EET strategies and may inform the design of more efficient bioelectrochemical systems.