Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.10.3.1 (tyrosinase)
9,065 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An enzyme carbon paste electrode containing three different enzymes was developed for the determination of L-phenylalanine. This sensor is based on the enzymatic/electrochemical recycling of tyrosinase in combination with salicylate hydroxylase and L-phenylalanine dehydrogenase (PADH). The enzymes salicylate hydroxylase and tyrosinase were coimmobilized first in a carbon paste electrode for the sensitive detection of NADH. The principle of the bienzyme scheme is as follows: the first enzyme, salicylate hydroxylase, converts salicylate to catechol in the presence of oxygen and NADH. The second enzyme, tyrosinase, then oxidizes the catechol to o-quinone, which is electrochemically detected and reduced back to catechol at the electrode at an Eappl = -50 mV vs Ag/AgCl. This results in an amplified signal due to the recycling of the catechol and o-quinone between tyrosinase and the surface of the electrode. Prior to adding PADH, the salicylate hydroxylase-tyrosinase carbon paste electrode was characterized in terms of its sensitivity to NADH, pH dependence, buffer composition, interferences, and stability. Interference from ascorbic acid and uric acid was found to be minimal. Human serum was used to investigate whether this bienzyme system was suitable for the detection of NADH in serum and blood samples. The sensitivity for NADH was increased by a factor of 33 times using the bienzyme amplification scheme (electroreduction of o-quinone at Eappl = -50 mV) as opposed to the salicylate hydroxylase single-enzyme system (at which catechol would have been oxidized at Eappl = +150 mV vs Ag/AgCl). The detection limit for NADH achieved by the bienzyme carbon paste electrode was 1 vs 100 microM for the single-enzyme carbon paste electrode. The salicylate hydroxylase-tyrosinase system was then coupled with phenylalanine dehydrogenase for L-phenylalanine determination. This multienzyme sensor was able to achieve a linear range of 20-150 microM and a detection limit of 5 microM for L-phenylalanine. The sensitivity is sufficient since the reference clinical range for L-phenylalanine is 78-206 microM.
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PMID:Determination of L-phenylalanine based on an NADH-detecting biosensor. 951 73

Dopamine has been implicated as a potential mediating factor in a variety of neurodegenerative disorders. Dopamine can be oxidized to form a reactive dopamine quinone that can covalently modify cellular macromolecules including protein and DNA. This oxidation can be enhanced through various enzymes including tyrosinase and/or prostaglandin H synthase. One of the potential targets in brain for dopamine quinone damage is tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis. The present studies demonstrated that dopamine quinone, the formation of which was enhanced through the activity of the melanin biosynthetic enzyme, tyrosinase, covalently modified and inactivated tyrosine hydroxylase. Dihydroxyphenylalanine (DOPA; the catechol-containing precursor of dopamine) also inactivated tyrosine hydroxylase under these conditions. Catecholamine-mediated inactivation occurred with both purified tyrosine hydroxylase as well as enzyme present in crude pheochromocytoma homogenates. Inactivation was associated with covalent incorporation of radiolabelled dopamine into the enzyme as assessed by immunoprecipitation, size exclusion chromatography, and denaturing sodium dodecylsulfate (SDS)-polyacrylamide gel electrophoresis. Furthermore, the covalent modification and inactivation of tyrosine hydroxylase was blocked by antioxidant compounds (dithiothreitol, reduced glutathione, or NADH). In addition to kinetic feedback inhibition and the formation of an inhibitory dopamine/Fe+3 complex, these findings suggest that a third mechanism exists by which dopamine (or DOPA) can inhibit tyrosine hydroxylase, adding further complexity to the regulation of catecholamine biosynthesis.
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PMID:Dopamine, in the presence of tyrosinase, covalently modifies and inactivates tyrosine hydroxylase. 984 60

A new spectrophotometric assay for the determination of the polyphenolic content of olive oil is presented. It is a substrate-recycling assay for phenolic compounds that employs tyrosinase in the presence of excess NADH. The reaction of various phenols with the enzyme produces an o-quinone, which is detected by recycling between reactions with the enzyme and NADH. The method offers some advantages over the classical methods employed to determine the polyphenolic content of olive oil, that is, ease and reproducibility of the analysis, highly increased sensitivity, and selectivity toward phenolic compounds. The amount of total polyphenols was determined in virgin olive oils both with the Folin-Ciocalteu reagent and with the proposed enzymatic method. The results suggest a better estimation of the polyphenol content, as compared with the colorimetric method. This has to be attributed to the different reactivities of the two methods toward phenols and catechols. Finally, the enzymatic method demonstrates that there is a linear relationship between the olive oil phenolic content and the antioxidative capacity of oil extracts.
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PMID:Enzymatic assay for the determination of olive oil polyphenol content: assay conditions and validation of the method. 1069 31

This work presents an amperometric biosensor incorporated into a flow configuration comprising salicylate hydroxylase that catalyses the irreversible hydroxylation of salicylate to catechol in the presence of NADH and molecular oxygen, and tyrosinase that further oxidises catechol giving o-quinone which is electrochemically reduced at -100 mV vs. Ag/AgCl yielding catechol and entering the catalytic oxidation and electrochemical reduction cycling which results in signal amplification and, consequently, low limits of detection. Additional incorporation of glucose dehydrogenase in the enzymatic sequence results in regeneration of NADH provided that glucose is present in the carrier stream and incorporation of a dialysis membrane provides operational stability to the biosensor. The analytical characteristics of this catalytic and electrochemical transduction sequence in a FIA system are: a limit of detection of 3.5 10(-6) M (S/N = 3), a sensitivity of 22.6 nA microM(-1) cm(-2), no loss of response at least after 5 h of continuous operation, and a sample frequency of 15 h(-1). Monitoring of salicylate after ingestion of 500 mg of acetylsalicylic acid has been followed in non-pretreated urine samples and the amount of salicylate in several drugs has been also successfully quantified.
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PMID:A new enzyme electrode for quantification of salicylic acid in a FIA system. 1069 72

Opiomelanins represent a new class of synthetic pigments produced by the tyrosinase-catalyzed oxidation of opioid peptides and other tyrosine aminoterminal peptides. In contrast with dopamelanin, these polymers are fully soluble in hydrophilic media, due to the presence of the peptide moiety. Opiomelanins show paramagnetism as demonstrated by the EPR spectrum identical to that of dopamelanin. The presence of the aminoacidic chain linked to the melaninic moiety, influences the electron transfer properties of opiomelanins i.e. the ability to oxidize NADH. Like dopamelanin Tyr-Gly-melanin exhibits this behaviour whereas leuenkmelanin does not show any oxidizing potential. Opiomelanins UV-Vis spectrum is characterized by an absorption band at 330 nm which disappears upon acid hydrolysis, H2O2 treatment and under simulated solar illumination. Opiomelanins exhibit a fluorescence emission peaked at 440 and 520 nm if excited at 330 nm. These fluorescence bands are due to the oligomeric units and high molecular weight units, respectively. When opioid peptides are allowed to react with tyrosinase in the presence of an excess of cysteine, cysteinyldopaenkephalins are synthesized. These peptides are furtherly oxidized giving rise to pheoopiomelanins. Reactive oxygen species also are able to oxidize non enzymatically both enkephalins and cysteinyldopaenkephalins, producing the corresponding melanin pigments.
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PMID:Opiomelanins synthesis and properties. 1151 Sep 84

A tyrosinase-directed therapeutic approach for treating malignant melanoma uses depigmenting phenolic prodrugs such as 4-hydroxyanisole (4-HA) for oxidation by melanoma tyrosinase to form cytotoxic o-quinones. However, in a recent clinical trial, both renal and hepatic toxicity were reported as side effects of 4-HA therapy. In the following, 4-HA (200 mg/kg i.p.) administered to mice caused a 7-fold increase in plasma transaminase toxicity, an indication of liver toxicity. Furthermore, 4-HA induced-cytotoxicity toward isolated hepatocytes was preceded by glutathione (GSH) depletion, which was prevented by cytochrome p450 inhibitors that also partly prevented cytotoxicity. The 4-HA metabolite formed by NADPH/microsomes and GSH was identified as a hydroquinone mono-glutathione conjugate. GSH-depleted hepatocytes were much more prone to cytotoxicity induced by 4-HA or its reactive metabolite hydroquinone (HQ). Dicumarol (an NAD(P)H/quinone oxidoreductase inhibitor) also potentiated 4-HA- or HQ-induced toxicity whereas sorbitol, an NADH-generating nutrient, prevented the cytotoxicity. Ethylenediamine (an o-quinone trap) did not prevent 4-HA-induced cytotoxicity, which suggests that the cytotoxicity was not caused by o-quinone as a result of 4-HA ring hydroxylation. Deferoxamine and the antioxidant pyrogallol/4-hydroxy-2,2,6,6-tetramethylpiperidene-1-oxyl (TEMPOL) did not prevent 4-HA-induced cytotoxicity, therefore excluding oxidative stress as a cytotoxic mechanism for 4-HA. A negligible amount of formaldehyde was formed when 4-HA was incubated with rat microsomal/NADPH. These results suggest that the 4-HA cytotoxic mechanism involves alkylation of cellular proteins by 4-HA epoxide or p-quinone rather than involving oxidative stress.
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PMID:Metabolic activation of 4-hydroxyanisole by isolated rat hepatocytes. 1222 81

A tyrosinase-directed therapeutic approach for malignant melanoma therapy uses the depigmenting phenolic agents such as 4-hydroxyanisole (4-HA) to form cytotoxic o-quinones. However, renal and hepatic toxicity was reported as side effects in a recent 4-HA clinical trial. In search of novel therapeutics, the cytotoxicity of the isomers 4-HA, 3-HA and 2-HA were investigated. In the following, the order of the HAs induced hepatotoxicity in mice, as measured by increased in vivo plasma transaminase activity, or in isolated rat hepatocytes, as measured by trypan blue exclusion, was 3-HA > 2-HA > 4-HA. Hepatocyte GSH depletion preceded HA induced cytotoxicity and a 4-MC-SG conjugate was identified by LC/MS/MS mass spectrometry analysis when 3-HA was incubated with NADPH/microsomes/GSH. 3-HA induced hepatocyte GSH depletion or GSH depletion when 3-HA was incubated with NADPH/microsomes was prevented by CYP 2E1 inhibitors. Dicumarol (an NAD(P)H: quinone oxidoreductase inhibitor) potentiated 3-HA- or 4-methoxycatechol (4-MC) induced toxicity whereas sorbitol (an NADH generating nutrient) greatly prevented cytotoxicity indicating a quinone-mediated cytotoxic mechanism. Ethylendiamine (an o-quinone trap) largely prevented 3-HA and 4-MC-induced cytotoxicity indicating that o-quinone was involved in cytotoxicity. Dithiothreitol (DTT) greatly reduced 3-HA and 4-MC induced toxicity. The ferric chelator deferoxamine slightly decreased 3-HA and 4-MC induced cytotoxicity whereas the antioxidants pyrogallol or TEMPOL greatly prevented the toxicity suggesting that oxidative stress contributed to 3-HA induced cytotoxicity. In summary, ring hydroxylation but not O-demethylation/epoxidation seems to be the bioactivation pathway for 3-HA in rat liver. The cytotoxic mechanism for 3-HA and its metabolite 4-MC likely consists cellular protein alkylation and oxidative stress. These results suggest that 3-HA is not suitable for treatment of melanoma.
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PMID:Metabolic activation of 3-hydroxyanisole by isolated rat hepatocytes. 1245 69

In the current work we investigated for the first time the biochemical basis of 4-hydroxyanisole (4-HA) induced toxicity in B16-F0 melanoma cells. It was found that dicoumarol, a diaphorase inhibitor, and 1-bromoheptane, a GSH depleting agent, increased 4-HA induced toxicity towards B16-F0 cells whereas dithiothreitol, a thiol containing agent, and ascorbic acid (AA), a reducing agent, largely prevented 4-HA toxicity. TEMPOL and pyrogallol, free radical scavengers, did not significantly prevent 4-HA toxicity towards B16-F0 cells. GSH>AA>NADH prevented the o-quinone formation when 4-HA was metabolized by tyrosinase/O(2). 4-HA metabolism by horseradish peroxidase/H(2)O(2) was prevented more effectively by AA than NADH>GSH. We therefore concluded that quinone formation was the major pathway for 4-HA induced toxicity in B16-F0 melanoma cells whereas free radical formation played a negligible role in the 4-HA induced toxicity.
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PMID:Biochemical basis of 4-hydroxyanisole induced cell toxicity towards B16-F0 melanoma cells. 1642 88

Ascorbic acid oxidase activity in Myrothecium verrucaria extracts resulted in O(2) uptake exceeding 0.5 mole per mole of ascorbic acid and in CO(2) evolution. Measurement of oxidized ascorbic acid at completion of the reaction demonstrated that an average of 10% of the oxidized product disappeared. A comparison of the gas exchange data with the amount of ascorbic acid not accounted for indicated that the reaction could not be explained by independent oxidase and oxygenase systems. Chromatographic examination of the reaction mixtures identified l-threonic acid. Experiments with ascorbic acid-1-(14)C showed that C-1 was partially decarboxylated during the oxidation. Test of the fungal extracts for enzymes that might explain the deviation from expected stoichiometry showed that phenolase, glutathione reductase, cytochrome oxidase, peroxidase and oxalic decarboxylase were not involved. Addition of azide in concentrations sufficient to block catalase increased excess O(2) consumption about 65%. No enzymes were found that could directly attack oxidized ascorbic acid. H(2)O(2) accumulated during oxidation in azide-blocked systems.The O(2) excess could be explained by assuming the enzyme had peroxidative capacity on a reductant other than ascorbic acid. An intermediate of ascorbic acid oxidation appeared to function as the substrate yielding CO(2) and l-threonic acid on degradation. The increase in excess O(2) utilized in azide-blocked systems and the H(2)O(2) accumulation also were explained by the proposed scheme.Another interpretation would involve production of free radicals during ascorbic acid oxidation. Evidence for this was the ability of extracts to oxidize DPNH in the presence of ascorbic acid. Oxygen radicals formed in such reactions were considered possible agents of degradation of ascorbic acid.
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PMID:Reaction Properties of the Ascorbic Acid Oxidase from Myrothecium verrucaria. 1665 40

We examined the ability of oxidation products of dopamine, DOPA, and 3,4-dihydroxyphenylacetic acid (DOPAC) to inhibit proteasomal activity. Dopamine, DOPA, and DOPAC underwent tyrosinase-catalyzed oxidation to generate aminochrome, dopachrome, and furanoquinone, respectively. In these studies, the oxidation of dopamine by tyrosinase generated product(s) that inhibited the proteasome, and proteasomal inhibition correlated with the presence of the UV-visible spectrum of aminochrome. The addition of superoxide dismutase and catalase did not prevent proteasomal inhibition. The addition of NADH and the quinone reductase NAD(P)H:quinone oxidoreductase 1 (NQO1) protected against aminochrome-induced proteasome inhibition. Although NQO1 protected against dopamine-induced proteasomal inhibition, the metabolism of aminochrome by NQO1 led to oxygen uptake because of the generation of a redox-labile cyclized hydroquinone, further demonstrating the lack of involvement of oxygen radicals in proteasomal inhibition. DOPA underwent tyrosinase-catalyzed oxidation to form dopachrome, and similar to aminochrome, proteasomal inhibition correlated with the presence of a dopachrome UV-visible spectrum. The inclusion of NQO1 did not protect against proteasomal inhibition induced by dopachrome. Oxidation of DOPAC by tyrosinase generated furanoquinone, which was a poor proteasome inhibitor. These studies demonstrate that oxidation products, including cyclized quinones derived from dopamine and related compounds, rather than oxygen radicals have the ability to inhibit the proteasome. They also suggest an important protective role for NQO1 in protecting against dopamine-induced proteasomal inhibition. The ability of endogenous intermediates formed during dopaminergic metabolism to cause proteasomal inhibition provides a potential basis for the selectivity of dopaminergic neuron damage in Parkinson's disease.
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PMID:A potential role for cyclized quinones derived from dopamine, DOPA, and 3,4-dihydroxyphenylacetic acid in proteasomal inhibition. 1679 May 33


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