Isolation, characterization, and primary structure of the vanadium chloroperoxidase from the fungus Embellisia didymospora

Here we describe the isolation, purification, and basic kinetic parameters of a vanadium type chloroperoxidase from the hyphomycete fungus Embellisia didymospora. The enzyme proved to possess similar high substrate affinities, a K-m of 5 mu M for a bromide, 1.2 mM for a chloride, and 60 mu M for a hydrogen peroxide, as those of the vanadium chloroperoxidase from Curvularia inaequalis, although with lower turnover rates for both Cl- and Br-. Substrate bromide was also found to inhibit the enzyme, a feature subsequently also noted for the chloroperoxidase from C, inaequalis, The gene encoding this enzyme was identified using DNA Southern blotting techniques and subsequently isolated and sequenced. A comparison is made between this vanadium chloroperoxidase and that of the fungus C, inaequalis both kinetically and at the sequence level. At the primary structural level the two chloroperoxidases share 68% identity, with conservation of all active site residues.