Water Oxidation by a Cytochrome P450: Mechanism and Function of the Reaction

P450_cam (CYP101A1) is a bacterial monooxygenase that is known to catalyze the oxidation of camphor, the first committed step in camphor degradation, with simultaneous reduction of oxygen (O₂). We report that P450_cam catalysis is controlled by oxygen levels: at high O₂ concentration, P450_cam catalyzes the known oxidation reaction, whereas at low O₂ concentration the enzyme catalyzes the reduction of camphor to borneol. We confirmed, using ¹⁷O and ²H NMR, that the hydrogen atom added to camphor comes from water, which is oxidized to hydrogen peroxide (H₂O₂). This is the first time a cytochrome P450 has been observed to catalyze oxidation of water to H₂O₂, a difficult reaction to catalyze due to its high barrier. The reduction of camphor and simultaneous oxidation of water are likely catalyzed by the iron-oxo intermediate of P450_cam, and we present a plausible mechanism that accounts for the 1∶1 borneol:H₂O₂ stoichiometry we observed. This reaction has an adaptive value to bacteria that express this camphor catabolism pathway, which requires O₂, for two reasons: 1) the borneol and H₂O₂ mixture generated is toxic to other bacteria and 2) borneol down-regulates the expression of P450_cam and its electron transfer partners. Since the reaction described here only occurs under low O₂ conditions, the down-regulation only occurs when O₂ is scarce.