The thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV oxidizes subatmospheric H2 with a high-affinity, membrane-associated [NiFe] hydrogenase

Abstract

The trace amounts (0.53 ppmv) of atmospheric hydrogen gas (H 2 ) can be utilized by microorganisms to persist during dormancy. This process is catalyzed by certain Actinobacteria, Acidobacteria, and Chloroflexi, and is estimated to convert 75 × 10 12 g H 2 annually, which is half of the total atmospheric H 2 . This rapid atmospheric H 2 turnover is hypothesized to be catalyzed by high-affinity [NiFe] hydrogenases. However, apparent high-affinity H 2 oxidation has only been shown in whole cells, rather than for the purified enzyme. Here, we show that the membrane-associated hydrogenase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV possesses a high apparent affinity (K m(app) = 140 nM) for H 2 and that methanotrophs can oxidize subatmospheric H 2 . Our findings add to the evidence that the group 1h [NiFe] hydrogenase is accountable for atmospheric H 2 oxidation and that it therefore could be a strong controlling factor in the global H 2 cycle. We show that the isolated enzyme possesses a lower affinity (K m = 300 nM) for H 2 than the membraneassociated enzyme. Hence, the membrane association seems essential for a high affinity for H 2 . The enzyme is extremely thermostable and remains folded up to 95°C. Strain SolV is the only known organism in which the group 1h [NiFe] hydrogenase is responsible for rapid growth on H 2 as sole energy source as well as oxidation of subatmospheric H 2 . The ability to conserve energy from H 2 could increase fitness of verrucomicrobial methanotrophs in geothermal ecosystems with varying CH 4 fluxes. We propose that H 2 oxidation can enhance growth of methanotrophs in aerated methane-driven ecosystems. Group 1h [NiFe] hydrogenases could therefore contribute to mitigation of global warming, since CH 4 is an important and extremely potent greenhouse gas.

A widely distributed hydrogenase oxidises atmospheric H2during bacterial growth

“…These findings contrast with multiple pure culture studies that have linked expression, activity, and phenotypes associated with group 1h [NiFe]-hydrogenases to survival rather than growth [10, 12, 18, 20, 22, 24, 25]. However, a growth-supporting role of atmospheric H 2 oxidation is nevertheless consistent with several surprising recent reports: the measurement of atmospheric H 2 oxidation during growth of several strains [12, 19, 24, 53]; the discovery of an Antarctic desert community driven by trace gas oxidation [9]; and the isolation of a proteobacterial methanotroph thought to grow on air alone [54]. Together, these findings suggest that the current persistence-centric model of atmospheric H 2 utilisation is overly generalised and that this process also supports growth.…”

“…Atmospheric H 2 oxidation is thought to be primarily mediated by group 1h [NiFe]-hydrogenases, a specialised oxygen-tolerant, high-affinity class of hydrogenases [4, 10–13]. To date, aerobic heterotrophic bacteria from four distinct bacterial phyla, the Actinobacteriota [10, 12, 14, 15], Acidobacteriota [16, 17], Chloroflexota [18], and Verrucomicrobiota [19], have been experimentally shown to consume atmospheric H 2 using this enzyme. This process has been primarily linked to energy conservation during persistence.…”

Structure of the 4-hydroxy-tetrahydrodipicolinate synthase from the thermoacidophilic methanotroph Methylacidiphilum fumariolicum SolV and the phylogeny of the aminotransferase pathway

“…Since M. fumariolicum SolV is a thermophile that can grow at temperatures of up to 65 C, it is conceivable that the enzymes in this bacterium are more thermostable compared with homologous enzymes in mesophilic microorganisms (Pol et al, 2007). This was recently shown for one of the hydrogenases encoded by M. fumariolicum SolV (Schmitz et al, 2020). Therefore, we searched for structural motifs that could enhance thermostability.…”

“…M. fumariolicum SolV was grown as a pure culture in a chemostat as described previously (Pol et al, 2007). Cell lysis and protein purification were performed as described previously (Schmitz et al, 2020). Briefly, the cell membranes were homogenized and gently stirred with 1%(w/v) n-dodecyl--d-maltoside for 1 h at room temperature.…”

“…Firstly, the solution was loaded onto a Q Sepharose column (GE Healthcare, Chicago, Illinois, USA) equilibrated with 20 mM bis-Tris pH 7.0. After washing with 20 mM bis-Tris, 100 mM NaCl pH 7.0 and with 20 mM bis-Tris, 200 mM NaCl pH 7.0, the most active fractions in terms of hydrogenase activity as determined by the method described by Schmitz et al (2020) The multistep reaction catalysed by DapA. The catalytic Lys162 is condensed with pyruvate, forming an enamine that reacts with l-aspartic semialdehyde (ASA).…”