Structure and mechanism of the epoxide hydrolase from Agrobacterium radiobacter AD1

The X-ray structure of the epoxide hydrolase from Agrobacterium radiobacter AD1 has been determined by isomorphous replacement at 2.1 Å resolution. The enzyme shows a two-domain structure, with the core having the α/β hydrolase fold topology which provides the scaffolding for the catalytic triad residues, Asp 107, Asp 246, and His 275. Based on biochemical and structural data, a mechanism is proposed to illuminate the peculiar chemical strategy to activate harmful epoxide substrates for hydrolysis and detoxification. The structure suggests Tyr 152/Tyr 215 as the residues involved in substrate binding, stabilization of the transition state, and possibly protonation of the epoxide oxygen. Site-directed mutagenesis studies confirmed the structural results and, in addition, showed a significant increase in enantioselectivity for styrene oxide, and substituted variants thereof, for a Tyr 215→Phe mutant. Since Tyr 215 is conserved among all epoxide hydrolase sequences, an activating role of this residue is suggested for other epoxide hydrolases as well, and mutation of this residue might also result in mutant enzymes with improved enantioselectivity in other cases.