Control of substrate access to the active site in methane monooxygenase

The crystal structure of the complex between the hydroxylase and regulatory component of soluble methane monooxygenase is presented, revealing how the latter component controls substrate access to the hydroxylase active site. Methanotrophs oxidize methane to methanol via soluble (sMMO) and particulate (pMMO) methane monooxygenases. sMMO comprises three protein components — a hydroxylase (MMOH), a reductase (MMOR), and a regulatory protein (MMOB) that couples electron consumption to substrate hydroxylation at the active site of MMOH. In this paper the authors solve the X-ray crystal structure of the MMOH–MMOB co-complex. The structure shows how MMOB controls the conformation of key amino acid residues in MMOH to enable the gaseous substrates to reach to its active site. Methanotrophs consume methane as their major carbon source and have an essential role in the global carbon cycle by limiting escape of this greenhouse gas to the atmosphere1,2,3. These bacteria oxidize methane to methanol by soluble and particulate methane monooxygenases (MMOs)1,2,3,4. Soluble MMO contains three protein components, a 251-kilodalton hydroxylase (MMOH), a 38.6-kilodalton reductase (MMOR), and a 15.9-kilodalton regulatory protein (MMOB), required to couple electron consumption with substrate hydroxylation at the catalytic diiron centre of MMOH2. Until now, the role of MMOB has remained ambiguous owing to a lack of atomic-level information about the MMOH–MMOB (hereafter termed H–B) complex. Here we remedy this deficiency by providing a crystal structure of H–B, which reveals the manner by which MMOB controls the conformation of residues in MMOH crucial for substrate access to the active site. MMOB docks at the α2β2 interface of α2β2γ2 MMOH, and triggers simultaneous conformational changes in the α-subunit that modulate oxygen and methane access as well as proton delivery to the diiron centre. Without such careful control by MMOB of these substrate routes to the diiron active site, the enzyme operates as an NADH oxidase rather than a monooxygenase5. Biological catalysis involving small substrates is often accomplished in nature by large proteins and protein complexes. The structure presented in this work provides an elegant example of this principle.