Crystal structure of a plant catechol oxidase containing a dicopper center

Abstract

Catechol oxidases are ubiquitous plant enzymes containing a dinuclear copper center. In the wound-response mechanism of the plant they catalyze the oxidation of a broad range of ortho-diphenols to the corresponding o-quinones coupled with the reduction of oxygen to water. The crystal structures of the enzyme from sweet potato in the resting dicupric Cu(II)-Cu(II) state, the reduced dicuprous Cu(I)-Cu(I) form, and in complex with the inhibitor phenylthiourea were analyzed. The catalytic copper center is accommodated in a central four-helix-bundle located in a hydrophobic pocket close to the surface. Both metal binding sites are composed of three histidine ligands. His 109, ligated to the CuA site, is covalently linked to Cys 92 by an unusual thioether bond. Based on biochemical, spectroscopic and the presented structural data, a catalytical mechanism is proposed in which one of the oxygen atoms of the diphenolic substrate binds to CuB of the oxygenated enzyme.

Tick attachment cement – reviewing the mysteries of a biological skin plug system

“…These coacervates release the glue, which gelates in the sea water because of the lower solubility of Ca/Mg phosphate, and irreversible cysteine–DOPA cross‐links are formed (dopaquinones). This cross‐linking is catalysed through monophenoloxidase and catechol oxidase activity of a tyrosinase, which is present in the secretory cells (Solomon, Sundaram & Machonkin, 1996; Klabunde et al ., 1998; Wang & Stewart, 2013). …”

Catecholase Activity of a Copper(II) Complex with a Macrocyclic Ligand: Unraveling Catalytic Mechanisms

“…Since the report on the crystal structure of catechol oxidase by Krebs and co-authors a few years ago, [1] the interest in model compounds of this biocatalyst, as well as for other type-3 copper enzymes (e.g., hemocyanin and tyrosinase) has taken a new turn. In particular, investigations on the substrate coordination to the copper centers, [2][3][4] the dioxygen activation and the subsequent reactivity of the peroxodicopper species, [5][6][7][8][9] and the role of a thioether linkage in a close proximity of the dicopper center, [10,11] found in catechol oxidase from Ipomoea batatas [1] and some hemocyanins [12,13] and tyrosinases, [14] have received a significant interest from the scientific community.…”

“…In particular, investigations on the substrate coordination to the copper centers, [2][3][4] the dioxygen activation and the subsequent reactivity of the peroxodicopper species, [5][6][7][8][9] and the role of a thioether linkage in a close proximity of the dicopper center, [10,11] found in catechol oxidase from Ipomoea batatas [1] and some hemocyanins [12,13] and tyrosinases, [14] have received a significant interest from the scientific community. Although there have been numerous reports on model complexes of catechol oxidase, detailed mechanistic studies on the catechol oxidation mechanism are unfortunately quite scarce, [15][16][17][18][19] and only in a few cases the mode of dioxygen reduction, for example, to dihydrogen peroxide or water, has been definitely established.…”

“…Although there have been numerous reports on model complexes of catechol oxidase, detailed mechanistic studies on the catechol oxidation mechanism are unfortunately quite scarce, [15][16][17][18][19] and only in a few cases the mode of dioxygen reduction, for example, to dihydrogen peroxide or water, has been definitely established. [15,16,18,[20][21][22][23][24] The mechanism of catechol oxidation by the natural enzyme, as proposed by Krebs and co-authors, [1] can be briefly outlined as follows. The native met (m-hydroxo dicopper(II)) form of the enzyme reacts stoichiometrically with one equivalent of the substrate, generating the corresponding quinone and the reduced deoxy (dicopper(I)) form.…”

“…The binding of catechol and dioxygen to the deoxy form may also occur simultaneously, as can be proposed from the crystal structure of the catechol oxidase adduct with the molecule of the inhibitor thiourea. [1] Recently, we reported the mechanism of catechol oxidation by a m-hydroxo-dicopper(II) complex with the macrocyclic ligand [22]py4pz (Figure 1, left). [8] In this complex, a single hydroxo bridge connects two copper(II) ions, each of which is further coordinated by four nitrogen donor atoms, provided by the ligand.…”