Component A2 is a redox-sensitive archaeal ATPase activated by methyl-coenzyme M reductase

Methyl-coenzyme M reductase (MCR) is the primary source of biogenic methane on Earth. In the active site of MCR, a nickel (Ni)-containing porphyrin (F430) must be in the Ni1+ oxidation state to initiate catalysis. The reductive activation of MCR, i.e., reduction of F430 to its Ni1+ state, is an ATP-dependent process, but the underlying ATPase and its precise role remain unknown. Component A2 is an ATP-binding protein that associates with MCR but, since it was reported to lack ATPase activity, its putative function was designated as an ATP-carrier protein. In contrast, recent structural insights into the MCR activation complex suggest that component A2 might hydrolyze ATP to drive conformational changes required for enzyme activation. Here, we provide direct biochemical evidence that component A2 is a bona fide ATPase that hydrolyzes ATP under strictly anaerobic conditions and only upon interaction with MCR. Mutational analyses reveal that component A2 must be bound to ATP prior to association with MCR and that residues involved in ATP hydrolysis do not impact protein-protein interaction. The two nucleotide-binding domains of A2 act cooperatively but display asymmetric contributions to ATP hydrolysis and MCR engagement. In addition, a distinctive N-terminal zinc-binding motif (ZBM) is required for maximal ATPase activity but is dispensable for MCR binding. Phylogenetic analyses reveal that this ZBM distinguishes component A2 from related ABC-type ATPases. Together, these findings identify component A2 as a distinct class of remodeling ATPases that powers conformational changes underlying the reductive activation of MCR. Significance StatementA large fraction of methane on Earth is generated by methanogenic archaea using the enzyme methyl coenzyme-M reductase (MCR). The maturation of MCR is a multi-step ATP-dependent process but the role of ATP and the corresponding ATPase(s) have remained unclear. Here, we show that component A2, a protein that is universally conserved in archaea that encode MCR and related enzymes, hydrolyzes ATP only upon interaction with MCR under anaerobic conditions. Our findings, together with recent structural studies, indicate that component A2-mediated ATP hydrolysis facilitates the reductive activation of MCR during the final step of its maturation. These results clarify a key step in the biogenesis of a central enzyme involved in biological methane production.