Comparative study of tyrosinases from different sources: Relationship between halide inhibition and the enzyme active site

Martinez, J.H.; Solano, Francisco; Peñafiel, Rafael; Galindo, Jesús D.; Iborra, José L.; Lozano, JoséA.

doi:10.1016/0305-0491(86)90309-3

Cited by 17 publications

(15 citation statements)

References 17 publications

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“…For bacterial Ty, we found that the apparent chloride inhibition constant (0.16 M at pH 6.80) is in the order of physiological chloride concentrations (5-200 mM). Similar values for the apparent chloride inhibition constant have been found for other Tys (17). A decrease in pH of 1 unit leads to a 10-fold increase in the fraction of Ty that is in the inactive chloride-bound form.…”

Section: Discussionsupporting

confidence: 67%

“…In an earlier study on Ty halide inhibition (17), different orders in halide inhibition strength were found for different tyrosinases (frog epidermis Ty,…”

Section: Discussionmentioning

confidence: 99%

“…The current knowledge concerning Ty halide inhibition is mainly limited to the observations that the inhibition is strongly pH-dependent and that the order of inhibition strength of F Ϫ , Cl Ϫ , Br Ϫ , and I Ϫ appears to be dependent on the source of the enzyme studied (16 -18). The latter differences have been explained mainly in terms of the accessibility of the halide ion to the Ty active site (17). Furthermore, EPR studies on halide-bound half-met hemocyanin derivatives have been reported (11,19), showing that halides interact directly with copper in the type-3 hemocyanin active site.…”

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confidence: 99%

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Structural Basis and Mechanism of the Inhibition of the Type-3 Copper Protein Tyrosinase from Streptomyces antibioticusby Halide Ions

Tepper¹,

Bubacco²,

Canters³

2002

Journal of Biological Chemistry

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The inhibition of the type-3 copper enzyme tyrosinase by halide ions was studied by kinetic and paramagnetic 1 H NMR methods. All halides are inhibitors in the conversion of L-3,4-dihydroxyphenylalanine (L-DOPA) with apparent inhibition constants that follow the order I ؊ < F ؊ < < Cl ؊ < Br ؊ at pH 6.80. The results show that the inhibition arises from the interaction of halide with both the oxidized (affinity F ؊ > Cl ؊ > Br ؊ > > I ؊ ) and reduced (affinity I ؊ > Br ؊ > Cl ؊ > > F ؊ ) enzyme. The paramagnetic 1 H NMR of the oxidized enzyme complexed with the halides is consistent with a direct interaction of halide with the type-3 site and shows that the (Cu-His 3 ) 2 coordination occurs in all halide-bound species. It is surmised that halides bridge both of the copper ions in the active site. Fluoride and chloride are shown to bind only to the low pH form of oxidized tyrosinase, explaining the strong pH dependence of the inhibition by these ions. We further show that p-toluic acid and the bidentate transition state analogue, Kojic acid, displace chloride from the oxidized active site, whereas the monodentate substrate analogue, p-nitrophenol, forms a ternary complex with the enzyme and the chloride ion. On the basis of the experimental results, a model is formulated for the inhibitor action and for the reaction of diphenols with the oxidized enzyme.One of the unresolved questions in the enzymology of the type-3 copper-containing tyrosinases (EC 1.14.18.1) is the detailed molecular mechanism of both inhibitor action and substrate conversion. This report focuses on the mechanism of their inhibition by halides. Tyrosinases are monooxygenating enzymes catalyzing the ortho-hydroxylation of monophenols and the subsequent oxidation of the diphenolic products to the corresponding quinones. The reactions take place under concomitant reduction of molecular oxygen to water. The formed quinones are reactive precursors in the synthesis of melanin pigments. In fruits, vegetables, and mushrooms, Ty 1 is a key enzyme in the browning that occurs upon bruising or long-term storage. In mammals, Ty is responsible for skin pigmentation. Defects in the enzyme may lead to some forms of oculocutaneous albinism or vitiligo (1). Furthermore, the enzyme has been linked to Parkinson's and other neurodegenerative diseases (2-6). Consequently, the enzyme poses considerable interest from medical, agricultural, and industrial points of view.The current knowledge of Ty at the biological, mechanistic, and structural levels has recently been reviewed (7-10). Ty harbors a dinuclear so-called type-3 copper center, the occurrence of which has also been established in hemocyanins, which act as oxygen carriers in arthropods and mollusks, and the catechol oxidases, which oxidize o-diphenols to the corresponding quinones. The two closely spaced copper ions in the type-3 active site are coordinated each by 3 histidine residues through the N⑀ nitrogen atoms (9). Although the known type-3 centers are found to be similar both in structure and in their a...

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Section: Discussionsupporting

confidence: 67%

“…In an earlier study on Ty halide inhibition (17), different orders in halide inhibition strength were found for different tyrosinases (frog epidermis Ty,…”

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Structural Basis and Mechanism of the Inhibition of the Type-3 Copper Protein Tyrosinase from Streptomyces antibioticusby Halide Ions

Tepper¹,

Bubacco²,

Canters³

2002

Journal of Biological Chemistry

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“…The Tyr-betaxanthin purified by FPLC had a high NaCl content that needed to be removed in order to avoid the inhibition of tyrosinase by chloride (Martínez et al, 1986). The high polarity of the pigments makes separation by organic solvents difficult.…”

Section: Extraction and Purification Of Dopaxanthin From Yellow Flowementioning

confidence: 99%

Betaxanthins as Substrates for Tyrosinase. An Approach to the Role of Tyrosinase in the Biosynthetic Pathway of Betalains

Escribano

García-Carmona

2005

Plant Physiology

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Tyrosinase or polyphenol oxidase (EC 1.14.18.1) is the key enzyme in melanin biosynthesis and in the enzymatic browning of fruits and vegetables. The role of tyrosinase in the secondary metabolism of plants still remains unclear, but its implication in betalain biosynthesis has been proposed. Betalains are an important class of water-soluble pigments, characteristic of plants belonging to the order Caryophyllales. In this article, the betaxanthins, tyrosine-betaxanthin (portulacaxanthin II) and dopaxanthin, are reported to be physiological substrates for tyrosinase. The direct activity of tyrosinase on selected betaxanthins is characterized in depth, and conversion of tyrosine-betaxanthin to dopaxanthin and its further oxidation to a series of compounds are described. Identity of the reaction products was studied by high-performance liquid chromatography and electrospray ionization-mass spectrometry. Masses determined for the reaction products were the same in all cases, 389 m/z ([M 1 H] 1 ) and equal to that determined for betanidin. Data indicate that dopaxanthin-quinone is obtained and evolves to more stable species by intramolecular cyclization. Kinetic parameters for tyrosinase acting on dopaxanthin were evaluated, showing a high affinity for this substrate (K m 5 84.3 mM). The biosynthetic scheme of betalains is reviewed and a branch is proposed based on the description of physiological substrates for tyrosinase. Lampranthus productus, Glottiphylum oligocarpum, and Glottiphylum pigmaeum are described as sources of stereopure (2S/S)-dopaxanthin.Tyrosinase or polyphenol oxidase (monophenol, o-diphenol:oxygen oxidoreductase; EC 1.14.18.1) is a copper enzyme that catalyzes two different reactions using molecular oxygen: the hydroxylation of monophenols to o-diphenols (monophenolase activity) and the oxidation of the o-diphenols to o-quinones (diphenolase activity; Sánchez-Ferrer et al., 1995). This enzyme is widely distributed in plants, microorganisms, and animals where tyrosinase is responsible for melanization.A wide variety of plant tyrosinase behaviors has been described and reviewed (Mayer and Harel, 1991;van Gelder et al., 1997). A molecular oxygenscavenging function in the chloroplast (Vaughn et al., 1988) and a role in plant defense (Constabel et al., 1995) have been suggested. Its implication in the secondary metabolism of betalains has been proposed (Steiner et al., 1999), but the physiological function of tyrosinase in higher plants is yet to be fully determined.Betalains are water-soluble nitrogen-containing pigments that are present in plants belonging to the order Caryophyllales (Strack et al., 2003) and in the fungal genera Amanita (Musso, 1979) and Hygrocybe (von Ardenne et al., 1974). All betalain pigments contain betalamic acid as the chromophore. Depending upon the nature of the betalamic acid addition residue, the betalains can be classified as either betacyanins or betaxanthins. Betacyanins contain a cyclo-3,4-dihydroxyphenylalanine (cyclo-DOPA; usually glycosylated) residue and exhibit ...

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“…The precursor amines were unable to interact with the column under the working conditions and were totally washed out as an unbound fraction (followed by absorbance at 280 nm). Afterwards, a C-18 cartridge was used again to remove the NaCl content, which is known to inhibit tyrosinase (Martı´nez et al 1986). Purity was confirmed by HPLC analysis with PDA detection.…”

Section: Chemical Synthesis and Purification Of Betaxanthins Derived mentioning

confidence: 99%

Characterization of the monophenolase activity of tyrosinase on betaxanthins: the tyramine-betaxanthin/dopamine-betaxanthin pair

Escribano

García-Carmona

2005

Planta

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Tyrosinase or polyphenol oxidase (EC 1.14.18.1) is the key enzyme responsible for melanin biosynthesis and for the enzymatic browning of fruits and vegetables. Although the function of tyrosinase in the secondary metabolism of plants remains unclear, it has been proposed that the enzyme plays a role in the betalain biosynthetic pathway. Betalains are an important class of water-soluble pigments, characteristic of plants belonging to the order Caryophyllales. In the present work, the betaxanthins tyramine-betaxanthin (miraxanthin III) and dopamine-betaxanthin (miraxanthin V) are reported as new natural substrates for tyrosinase. The result of the diphenolase activity of the enzyme on dopamine-betaxanthin was a series of products identified by HPLC and ESI-MS as quinone-derivatives. Data indicate that dopamine-betaxanthin-quinone is obtained and evolves to more stable species by intramolecular cyclization. The kinetic parameters evaluated for the diphenolase activity were V(m) = 74.4 microM min(-1), K(m) = 94.7 microM. Monophenolase activity on tyramine-betaxanthin yielded the same compounds in the absence of a reducing agent, but when ascorbic acid was present enzymatic conversion to dopamine-betaxanthin could be found. For the first time, kinetic characterization of the monophenolase activity of tyrosinase on betaxanthins is provided (V(m) = 10.4 microM min(-1) and K(m) = 126.9 microM) and a lag period is described and analyzed according to the mechanism of action of the enzyme. The high affinity shown by tyrosinase for these substrates may be indicative of a previously unconsidered physiological role in betalain metabolism. A possible mechanism for the formation of 2-descarboxy-betacyanins from tyramine-betaxanthin by tyrosinase is also discussed.

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Comparative study of tyrosinases from different sources: Relationship between halide inhibition and the enzyme active site

Cited by 17 publications

References 17 publications

Structural Basis and Mechanism of the Inhibition of the Type-3 Copper Protein Tyrosinase from Streptomyces antibioticusby Halide Ions

Structural Basis and Mechanism of the Inhibition of the Type-3 Copper Protein Tyrosinase from Streptomyces antibioticusby Halide Ions

Betaxanthins as Substrates for Tyrosinase. An Approach to the Role of Tyrosinase in the Biosynthetic Pathway of Betalains

Characterization of the monophenolase activity of tyrosinase on betaxanthins: the tyramine-betaxanthin/dopamine-betaxanthin pair

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