Carbonic anhydrase plays multiple roles in acetotrophic growth of a model marine methanogen from the domainArchaea

Abstract

AbstractCarbonic anhydrase (CA) catalyzes the reversible hydration of CO2to bicarbonate and a proton. The enzyme is universally distributed in all three domains of life and plays diverse physiological roles in the domainsEukaryaandBacteria. Remarkably, a physiological role has not been identified for any CA from the domainArchaea. Herein are described roles for a gamma class CA (Cam) from the methane-producing marine archaeonMethanosarcina acetivorans. Acetate-dependent growth of a Δcammutant showed an extended lag phase, lower final cell density, and metabolized acetate to a threshold of 20.0 mM compared to 1.0 mM for wild-type. Molar growth yields (Ymethane) were substantially greater for wild-type compared to the mutant. In contrast, growth parameters were identical for the methanol-grown wild-type and mutant. Rates of methane formation in resting cell suspensions containing 20.0 mM acetate were significantly less in the mutantversuswild-type and dependent on the presence of CO2. Rates for the wild-type decreased with increasing pH that was more pronounced for the mutant. CA activity was 100-fold greater in the membraneversussoluble fraction of acetate-grown cells. Addition of a surrogate CA stimulated acetate-dependent methanogenesis in resting cell suspensions of the mutant. The results support a role for Cam to supply protons for symport of acetate by the AceP symporter that also optimizes and facilitates growth at low acetate concentrations and high pH values encountered in the marine environment whereM. acetivoranswas isolated.Significance StatementAlthough CA plays major physiological roles in the domainsEukaryaandBacteria, a role has not been reported for the domainArchaeain which methanogens comprise the major group with abundant genomic annotations for CAs. Acetotrophic methanogens account for most of the methane produced in Earth’s biosphere where it is a major greenhouse gas. Although the biochemistry of the conversion of acetate to methane and carbon dioxide is well known, little is understood of acetate transport. The finding that CA has multiple roles facilitating thermodynamically constrained growth of a model marine acetotrophic methanogen has implications for advancing ecological understanding of the methane cycle that impacts global warming and climate change. Finally, the work is an introduction to anticipated physiological roles of CAs in the domainArchaeafor which genomic annotations are abundant.