The vanadium chloroperoxidase from the fungus, <i>Curvularia inaequalis</i>

Schijndel, J.W.P.M. van; Simons, L.H.; Vollenbroek, Esther G. M.; Wever, R.

doi:10.1016/0014-5793(93)80811-8

Cited by 29 publications

(23 citation statements)

References 19 publications

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“…Incubation of fully activated r-CPO with p-NPP at pH 7.0 did not result in p-NPP hydrolysis while incubation at low pH did (results not shown). This is in agreement with results obtained earlier by our group (30), showing that that the K d of V-CPO for vanadate increases strongly at low pH, thus enabling p-NPP to compete effectively for the binding site. Fig.…”

Section: Resultssupporting

confidence: 83%

From phosphatases to vanadium peroxidases: A similar architecture of the active site

Hemrika¹,

Renirie²,

Dekker³

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

173

164

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We show here that the amino acid residues contributing to the active sites of the vanadate containing haloperoxidases are conserved within three families of acid phosphatases; this suggests that the active sites of these enzymes are very similar. This is confirmed by activity measurements showing that apochloroperoxidase exhibits phosphatase activity. These observations not only reveal interesting evolutionary relationships between these groups of enzymes but may also have important implications for the research on acid phosphatases, especially glucose-6-phosphatase-the enzyme affected in von Gierke disease-of which the predicted membrane topology may have to be reconsidered.Haloperoxidases are enzymes catalyzing the two electron oxidation of a halide (X Ϫ ) to the corresponding hypohalous acid according to Eq. 1.HOX may further react with a broad range of nucleophilic acceptors to form a diversity of halogenated compounds. These haloperoxidases are named after the most electronegative halide they are able to oxidize, and thus a chloroperoxidase (CPO) is able to oxidize chloride, bromide, and iodide. Three classes of haloperoxidases have been identified. One of these consists of enzymes without a prosthetic group and as such have been detected in a number of bacteria (1, 2). The remaining two classes are the so-called heme-containing haloperoxidases-exemplified by the CPO from the fungus Caldariomyces fumago (3) or myeloperoxidase which is present in white blood cells (4)-and the vanadium-containing haloperoxidases that bind a vanadate ion (VO 4 3Ϫ ) as a prosthetic group. Enzymes representing these two classes not only differ in the nature of their prosthetic group but also in at least two other aspects: catalytic mechanism and stability. Hemecontaining peroxidases catalyze the formation of the hypohalous acid by a redox mechanism, whereas in vanadatecontaining peroxidases the transition metal does not change its redox state (5, 6) but may function as a Lewis acid. Vanadatecontaining haloperoxidases not only posses a very high stability (7, 8) but they also resist a high concentration of their substrate (H 2 O 2 ) (9) and their product (HOX) (7) that would readily inactivate the heme-containing peroxidases.Vanadate-containing peroxidases were first discovered in seaweeds about a decade ago (10, 11); at present they are being discovered not only in an increasing number of seaweeds (12, 13) but also in fungi (8) and in a lichen (14).The fungus Curvularia inaequalis has been shown to express a vanadium-containing CPO (V-CPO) (15), and recently the gene encoding this enzyme was cloned and sequenced (16). The V-CPO gene codes for a protein of 609 amino acids with a calculated molecular mass of 67,488 kDa. Database searches using the entire V-CPO sequence showed very little sequence similarity to other known proteins (16). The recent determination of the crystal structure of this enzyme at 2.1-Å resolution (17) revealed that the protein has an overall cylindrical shape and measures approximately 50 ϫ 80 Å. Th...

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Section: Resultssupporting

confidence: 83%

From phosphatases to vanadium peroxidases: A similar architecture of the active site

Hemrika¹,

Renirie²,

Dekker³

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

173

164

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“…3B using a vanadate concentration range of 0 -500 M resulted in an almost linear relationship between the absorbance and vanadate concentration (not shown), with an intensity approximately 2.5 times higher than that of free vanadate at this pH. This indicates that the mutant has a lower affinity for vanadate at pH 5.0 than at pH 8.3, a phenomenon that has also been reported for the wild type enzyme (20).…”

Section: Resultsmentioning

confidence: 50%

Peroxidase and Phosphatase Activity of Active-site Mutants of Vanadium Chloroperoxidase from the Fungus Curvularia inaequalis

Renirie

Hemrika

Wever

2000

Journal of Biological Chemistry

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Mutation studies were performed on active-site residues of vanadium chloroperoxidase from the fungus Curvularia inaequalis, an enzyme which exhibits both haloperoxidase and phosphatase activity and is related to glucose-6-phosphatase. The effects of mutation to alanine on haloperoxidase activity were studied for the proposed catalytic residue His-404 and for residue Asp-292, which is located close to the vanadate cofactor. The mutants were strongly impaired in their ability to oxidize chloride but still oxidized bromide, although they inactivate during turnover. The effects on the optical absorption spectrum of vanadium chloroperoxidase indicate that mutant H404A has a reduced affinity for the cofactor, whereas this affinity is unchanged in mutant D292A. The effect on the phosphatase activity of the apoenzyme was investigated for six mutants of putative catalytic residues. Effects of mutation of His-496, Arg-490, Arg-360, Lys-353, and His-404 to alanine are in line with their proposed roles in nucleophilic attack, transition-state stabilization, and leaving-group protonation. Asp-292 is excluded as the group that protonates the leaving group. A model based on the mutagenesis studies is presented and may serve as a template for glucose-6-phosphatase and other related phosphatases. Hydrolysis of a phospho-histidine intermediate is the ratedetermining step in the phosphatase activity of apochloroperoxidase, as shown by burst kinetics.

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“…These fungi produce extracellular chloroperoxidases (CPOs), heme-or vanadium-containing enzymes that oxidize Cl Ϫ to hypochlorous acid (HOCl) or a similarly reactive chlorine electrophile (18,21). CPOs chlorinate a variety of aromatic substrates (2,15,19,23), but their reactivity with lignin is unknown.…”

mentioning

confidence: 99%

Chlorination and Cleavage of Lignin Structures by Fungal Chloroperoxidases

Ortiz-Bermúdez

Srebotnik

Hammel

2003

Appl Environ Microbiol

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Two fungal chloroperoxidases (CPOs), the heme enzyme from Caldariomyces fumago and the vanadium enzyme from Curvularia inaequalis, chlorinated 1-(4-ethoxy-3-methoxyphenyl)-2-(2-methoxyphenoxy)-1,3-dihydroxypropane, a dimeric model compound that represents the major nonphenolic structure in lignin. Both enzymes also cleaved this dimer to give 1-chloro-4-ethoxy-3-methoxybenzene and 1,2-dichloro-4-ethoxy-5-methoxybenzene, and they depolymerized a synthetic guaiacyl lignin. Since fungal CPOs occur in soils and the fungi that produce them are common inhabitants of plant debris, CPOs may have roles in the natural production of high-molecular-weight chloroaromatics and in lignin breakdown.Natural organochlorine compounds are widely distributed in the environment, but the processes that generate them are not completely understood. It is clear that many natural organochlorines are microbial secondary metabolites, whereas others are generated abiotically (22), but little attention has been given to the possibility that biological chlorinations of lignin, the most abundant terrestrial aromatic substance, may produce chloroaromatic compounds. If microbial mechanisms exist to oxidize Cl Ϫ in plant material, the resulting electrophilic chlorine species are likely to react with the electron-rich aromatic rings of lignin (4). This chemistry could explain, in part, why field samples of soil, litter, and decayed wood have all been shown to contain high-molecular-weight chloroaromatics (5,14).Certain plant-pathogenic ascomycetes that are cosmopolitan inhabitants of plant debris provide a possible route for Cl Ϫ oxidation in the vicinity of lignin (6,8,20). These fungi produce extracellular chloroperoxidases (CPOs), heme-or vanadium-containing enzymes that oxidize Cl Ϫ to hypochlorous acid (HOCl) or a similarly reactive chlorine electrophile (18,21). CPOs chlorinate a variety of aromatic substrates (2,15,19,23), but their reactivity with lignin is unknown. We report here that the major structures in lignin are not only chlorinated but also cleaved by the heme CPO of Caldariomyces fumago and the vanadium CPO of Curvularia inaequalis.Chlorination and cleavage of a lignin model dimer. Compound I, 14 C]phenyl)-2-(2-methoxyphenoxy)-1,3-dihydroxypropane (1.0 mCi/mmol), a model of the major nonphenolic arylglycerol-␤-aryl ether structure in lignin (Fig. 1A), was prepared as described previously (9, 12). The model was treated with either commercially available Caldariomyces fumago heme CPO (Sigma) or partially purified Curvularia inaequalis vanadium CPO. The latter enzyme was obtained by DEAE-Sephacel chromatography of the extracellular medium from Curvularia inaequalis cultures (ATCC 10713) as described earlier (17). The reaction mixtures (1.0 ml) contained compound I (3.4 ϫ 10 5 dpm, 160 M), KCl (20 mM), and buffer (25 mM). The buffers were potassium phosphate, pH 3.0, for heme CPO reactions and sodium acetate, pH 4.0, for vanadium CPO reactions. Enzyme was added (35 monochlorodimedone units of heme CPO or 1 monochlorodimedone unit of va...

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The vanadium chloroperoxidase from the fungus, Curvularia inaequalis

Cited by 29 publications

References 19 publications

From phosphatases to vanadium peroxidases: A similar architecture of the active site

From phosphatases to vanadium peroxidases: A similar architecture of the active site

Peroxidase and Phosphatase Activity of Active-site Mutants of Vanadium Chloroperoxidase from the Fungus Curvularia inaequalis

Chlorination and Cleavage of Lignin Structures by Fungal Chloroperoxidases

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