Spectroscopic and substrate binding properties of heme-containing aldoxime dehydratases, OxdB and OxdRE

Aldoxime dehydratase (Oxd) is a novel hemeprotein that catalyzes the dehydration reaction of aldoxime to produce nitrile. In this study, we studied the spectroscopic and substrate binding properties of two Oxds, OxdB from Bacillus sp. strain OxB-1 and OxdRE from Rhodococcus sp. N-771, that show different quaternary structures and relatively low amino acid sequence identity. Electronic absorption and resonance Raman spectroscopy revealed that ferric OxdRE contained a six-coordinate low-spin heme, while ferric OxdB contained a six-coordinate high-spin heme. Both ferrous OxdRE and OxdB included a five-coordinate high-spin heme to which the substrate was bound via its nitrogen atom for the reaction to occur. Although the ferric Oxds were inactive for catalysis, the substrate was bound to the ferric heme via its oxygen atom in both OxdB and OxdRE. Electronic paramagnetic resonance (EPR) and rapid scanning spectroscopy revealed that the flexibility of the heme pocket was different between OxdB and OxdRE, which might affect their substrate specificity.