Unification for the Expression of the Monophenolase and Diphenolase Activities of Tyrosinase

Fenoll, Lorena G.; Rodrı́guez-López, José Neptuno; García‐Molina, Francisco; Garcı́a-Cánovas, Francisco; Tudela, José

doi:10.1080/15216540214537

Cited by 15 publications

(11 citation statements)

References 21 publications

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“…One unit of PO specific activity corresponds to the amount of enzyme that catalyzes the production of 1 µmol of dopachrome ig. 1 Estuarine sampling areas in French Atlantic coast: a localization of reference (R) and contaminated (C) sites in France and b localization of contaminated sites Mus du Loup (C1), Boyard (C2), and Les Palles (C3) in Marennes-Oleron Bay (ε Dopachrome =3,300 M −1 cm −1 ) per minute (Pomerantz 1963;Fenoll et al 2002).…”

Section: Biochemical Analysismentioning

confidence: 99%

Seasonal variation of pollution biomarkers to assess the impact on the health status of juvenile Pacific oysters Crassostrea gigas exposed in situ

Luna‐Acosta

Bustamante

Godefroy

et al. 2010

Environ Sci Pollut Res

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Section: Biochemical Analysismentioning

confidence: 99%

Seasonal variation of pollution biomarkers to assess the impact on the health status of juvenile Pacific oysters Crassostrea gigas exposed in situ

Luna‐Acosta

Bustamante

Godefroy

et al. 2010

Environ Sci Pollut Res

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“…Specific activities (SA) were expressed in international units (IU) per mg of total protein. One IU is defined as the amount of enzyme that catalyzes the appearance of 1 mmol of product per min [24]. Apparent MichaeliseMenten constants (Km app ) and maximum velocities (Vm app ) were estimated from double reciprocal plots (LineweavereBurk) of velocity vs substrate concentration.…”

Section: Phenoloxidase Assaysmentioning

confidence: 99%

First evidence of laccase activity in the Pacific oyster Crassostrea gigas

Luna‐Acosta¹,

Rosenfeld²,

Amari³

et al. 2010

Fish & Shellfish Immunology

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Phenoloxidases (POs) are a family of enzymes including tyrosinases, catecholases and laccases, which play an important role in immune defence mechanisms in various invertebrates. The aim of this study was to thoroughly identify the PO-like activity present in the hemolymph of the Pacific oyster Crassostrea gigas, by using different substrates (i.e. dopamine and p-phenylenediamine, PPD) and different PO inhibitors. In order to go deeper in this analysis, we considered separately plasma and hemocyte lysate supernatant (HLS). In crude plasma, oxygraphic assays confirmed the presence of true oxidase activities. Moreover, the involvement of peroxidase(s) was excluded. In contrast to other molluscs, no tyrosinase-like activity was detected. With dopamine as substrate, PO-like activity was inhibited by the PO inhibitors tropolone, phenylthiourea (PTU), salicylhydroxamic acid and diethyldithio-carbamic acid, by a specific inhibitor of tyrosinases and catecholases, i.e. 4-hexylresorcinol (4-HR), and by a specific inhibitor of laccases, i.e. cetyltrimethylammonium bromide (CTAB). With PPD as substrate, PO-like activity was inhibited by PTU and CTAB. In precipitated protein fractions from plasma, and with dopamine and PPD as substrates, PTU and 4-HR, and PTU and CTAB inhibited PO-like activity, respectively. In precipitated protein fractions from hemocyte lysate supernatant, PTU and CTAB inhibited PO-like activity, independently of the substrate. Taken together, these results suggest the presence of both catecholase- and laccase-like activities in plasma, and the presence of a laccase-like activity in HLS. To the best of our knowledge, this is the first time that a laccase-like activity is identified in a mollusc by using specific substrates and inhibitors for laccase, opening new perspectives for studying the implication of this enzyme in immune defence mechanisms of molluscs of high economic value such as C. gigas.

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“…One IU is defined as the amount of enzyme that catalyses the appearance of 1 mmole of product per min [30], using molar extinction coefficients of L-DOPA and PPD reaction products of 3600 M À1 cm À1 [31] and 43 160 M À1 cm À1 [32,33] under these conditions, respectively. The protein concentration of oyster tissue extracts was determined by the slightly modified Lowry method, as described previously [34].…”

Section: Phenoloxidase Assaysmentioning

confidence: 99%

First evidence of a potential antibacterial activity involving a laccase-type enzyme of the phenoloxidase system in Pacific oyster Crassostrea gigas haemocytes

Luna‐Acosta

Saulnier

Pommier

et al. 2011

Fish & Shellfish Immunology

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Phenoloxidases (POs) are a group of copper proteins including tyrosinase, catecholase and laccase. In several insects and crustaceans, antibacterial substances are produced through the PO cascade, participating in the direct killing of invading microorganisms. However, although POs are widely recognised as an integral part of the invertebrate immune defence system, experimental evidence is lacking that these properties are conserved in molluscs, and more particularly in the Pacific oyster Crassostrea gigas. In the present study, Vibrio splendidus LGP32 and Vibrio aestuarianus 02/041 growths were affected, after being treated with C. gigas haemocyte lysate supernatant (HLS), and either a common substrate of POs, l-3,4-dihydroxyphenylalanine (L-DOPA), to detect catecholase-type PO activity, or a specific substrate of laccase, p-phenylenediamine (PPD), to detect laccase-type PO activity. Interestingly, a higher bacterial growth inhibition was observed in the presence of PPD than in the presence of L-DOPA. These effects were suppressed when the specific PO inhibitor, phenylthiourea (PTU), was added to the medium. Results of the present study suggest, for the first time in a mollusc species, that antibacterial activities of HLS from C. gigas potentially involve POs, and more particularly laccase catalysed reactions.

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Unification for the Expression of the Monophenolase and Diphenolase Activities of Tyrosinase

Cited by 15 publications

References 21 publications

Seasonal variation of pollution biomarkers to assess the impact on the health status of juvenile Pacific oysters Crassostrea gigas exposed in situ

Seasonal variation of pollution biomarkers to assess the impact on the health status of juvenile Pacific oysters Crassostrea gigas exposed in situ

First evidence of laccase activity in the Pacific oyster Crassostrea gigas

First evidence of a potential antibacterial activity involving a laccase-type enzyme of the phenoloxidase system in Pacific oyster Crassostrea gigas haemocytes

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