Mechanistic Investigation of a Non-Heme Iron Enzyme Catalyzed Epoxidation in (−)-4′-Methoxycyclopenin Biosynthesis

Mechanisms have been proposed for α-KG-dependent non-heme iron enzyme catalyzed oxygen atom insertion into an olefinic moiety in various natural products, but they have not been examined in detail. Using a combination of methods including transient kinetics, Mössbauer spectroscopy, and mass spectrometry, we demonstrate that AsqJ-catalyzed (−)-4′-methoxy­cyclopenin formation uses a high-spin Fe­(IV)-oxo intermediate to carry out epoxidation. Furthermore, product analysis on 16O/18O isotope incorporation from the reactions using the native substrate, 4′-methoxy­dehydro­cyclopeptin, and a mechanistic probe, dehydro­cyclopeptin, reveals evidence supporting oxo↔hydroxo tautomerism of the Fe­(IV)-oxo species in the non-heme iron enzyme catalysis.