Particulate methane monooxygenase (pMMO) is an integral membrane metalloenzyme that converts methane to methanol in methanotrophic bacteria. The enzyme consists of three subunits, pmoB, pmoA, and pmoC, organized in an α 3β 3γ 3 trimer. Studies of intact pMMO and a recombinant soluble fragment of the pmoB subunit (denoted as spmoB) indicate that the active site is located within the soluble region of pmoB at the site of a crystallographically modeled dicopper center. In this work, we have investigated the reactivity of pMMO and spmoB with oxidants. Upon reduction and treatment of spmoB with O 2 or H 2O 2 or pMMO with H 2O 2, an absorbance feature at 345 nm is generated. The energy and intensity of this band are similar to those of the μ-η 2:η 2-peroxo-Cu II 2 species formed in several dicopper enzymes and model compounds. The feature is not observed in inactive spmoB variants in which the dicopper center is disrupted, consistent with O 2 binding to the proposed active site. Reaction of the 345 nm species with CH 4 results in the disappearance of the spectroscopic feature, suggesting that this O 2 intermediate is mechanistically relevant. Taken together, these observations provide strong new support for the identity and location of the pMMO active site.
ASJC Scopus subject areas
- Colloid and Surface Chemistry