X-ray structure determination of a vanadium-dependent haloperoxidase from Ascophyllum nodosum at 2.0 angstrom resolution

The homo-dimeric structure of a vanadium-dependent haloperoxidase (V-BPO) f rom the brown alga Ascophyllum nodosum (EC 1.1.11.X) has been solved by sin gle isomorphous replacement anomalous scattering (SIRAS) X-ray crystallogra phy at 2.0 Angstrom resolution (PDB accession code 1QI9), using two heavy-a tom datasets of a tungstate derivative measured at two different wavelength s. The protein sequence (SwissProt entry code P81701) of V-BPO was establis hed by combining results from protein and DNA sequencing, and electron dens ity interpretation. The enzyme has nearly an all-helical structure, with tw o four-helix bundles and only three small beta-sheets. The holoenzyme conta ins trigonal-bipyramidal coordinated vanadium atoms at its two active centr es. Structural similarity to the only other structurally characterized vana dium-dependent chloroperoxidase (V-CPO) from Curvularia inaequalis exists i n the vicinity of the active site and to a lesser extent in the central fou r-helix bundle. Despite the low sequence and structural similarity between V-BPO and V-CPO, the vanadium binding centres are highly conserved on the N-terminal side o f an alpha-helix and include the proposed catalytic histidine residue (His4 18(V-BPO)/His(V-CPO)). The V-BPO structure contains, in addition, a second histidine near the active site (His411(V-BPO)), which can alter the redox p otential of the catalytically active VO2-O-2 species by protonation/deproto nation reactions. Specific binding sites for the organic substrates, like i ndoles and monochlordimedone, or for halide ions are not visible in the V-B PO structure. A reaction mechanism for the enzymatic oxidation of halides i s discussed, based on the present structural, spectroscopic and biochemical knowledge of vanadium-dependent haloperoxidases, explaining the observed e nzymatic differences between both enzymes. (C) 1999 Academic Press.