Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.10.3.1 (tyrosinase)
 9,065 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe a convenient and sensitive assay of polyphenol oxidase (PPO, EC 1.14.18.1) consisting of spectrophotometry at 300 nm based on the stoichiometric reaction of cysteine with o-quinones produced during the enzymatic oxidation of phenols. The adduct formed exhibited spectral properties different from those of the parent phenol. PPO activities extracted from apple, pear, and mushroom were assayed. The assay cannot be used with hydroxycinnamoyl derivatives since the cysteinyl adduct compounds exhibited spectral properties similar to those of their parent phenolic substrates. However, the cysteine-coupled method presents several advantages over the measurement of oxygen uptake by polarography or the direct estimation of o-quinone by spectrophotometry. The duration of the linear period was increased, allowing a better estimation of its value. The zone of proportionality between rates and enzyme quantities was enlarged. The difference in molar extinction coefficients between adduct and phenol at 300 nm ranged between 2000 and 2800 M-1.cm-1, i.e., two times higher than those of the corresponding o-quinones. Therefore, this assay improves the sensitivity of polyphenol oxidase detection over that of the direct spectrophotometric assay of quinone formation.
 ...
PMID:New spectrophotometric assay for polyphenol oxidase activity. 829 16

For the first time, unfolding (6 M guanidine) and refolding of partially proteolysed purified polyphenol oxidase (PPOr) was achieved, with 88% of activity recovered. Optimal refolding conditions consisted in stepwise dialysis of guanidine treated extracts, the dialysis buffers containing 1 M (NH4)2SO4 and 100 microM CuSO4. However, CuSO4 had limited effect on the recovering of PPOr activity, whereas (NH4)2SO4 was essential. Concerning the PPO tertiary structure, denaturing conditions (combinations of boiling and reducing agent) used on SDS-PAGE have shown (i) a compact tertiary structure and (ii) the presence of disulfide bonds in PPOr, accounting for the shift between 27 and 41 kDa, and 41 and 42 kDa, respectively. Resistance to proteolytic cleavage was used to study the conformational changes induced by the denaturing treatments. Folded PPOr was resistant to further proteolysis whereas unfolded PPO was totally digested, indicating the role of tertiary structure of PPOr in the resistance to proteases.
 ...
PMID:Unfolding and refolding of active apple polyphenol oxidase. 984 26

The oxidative end products that result from the biocatalysis of tyrosinase (PPO) and/or a polyphenol esterase (PPE) extract have been investigated simultaneously in model systems containing selected phenolic compounds as substrates. The spectrophotometric scanning of brown color, formed in the presence of both PPO and PPE, showed a decrease in the absorbance compared to that obtained with PPO only. Graphical analyses of the iterative spectra of oxidized phenolic end products by PPO confirmed the presence of, at least, three kinetically related absorbing species. HPLC analyses of the end products, obtained by the biocatalysis of PPE or PPO activity, indicated the presence of two main groups of compounds: colored ones of lambda(max) at 294-324 nm and colorless products of lambda(max) at 264-290 nm. PPE produced both compounds when selected substrates were used as substrates, whereas PPO produced only one type of oxidation product. However, when both enzymes were incubated together, the nature of the end products was similar to that obtained with PPE only.
 ...
PMID:Characterization of tyrosinase- and polyphenol esterase-catalyzed end products using selected phenolic substrates. 1079 54

Polyphenol oxidase (EC 1.10.3.1, PPO) in the pulp of banana (Musa sapientum L.) was purified to 636-fold with a recovery of 3.0%, using dopamine as substrate. The purified enzyme exhibited a clear single band on polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate (SDS)-PAGE. The molecular weight of the enzyme was estimated to be about 41000 and 42000 by gel filtration and SDS-PAGE, respectively. The enzyme quickly oxidized dopamine, and its K(m) value for dopamine was 2.8 mM. The optimum pH was at 6.5, and the enzyme activity was stable in the range of pH 5-11 at 5 degrees C for 48 h. The enzyme had an optimum temperature of 30 degrees C and was stable even after a heat treatment at 70 degrees C for 30 min. The enzyme activity was completely inhibited by L-ascorbic acid, cysteine, sodium diethyldithiocarbamate, and potassium cyanide. Under a low buffer capacity, the enzyme was also strongly inhibited by citric acid and acetic acid at 10 mM.
 ...
PMID:Purification and characterization of polyphenol oxidase from banana (Musa sapientum L.) pulp. 1089 14

Polyphenol oxidase (EC 1.10.3.1, o-diphenol: oxygen oxidoreductase, PPO) of banana (Musa sapientum L.) peel was partially purified about 460-fold with a recovery of 2.2% using dopamine as substrate. The enzyme showed a single peak on Toyopearl HW55-S chromatography. However, two bands were detected by staining with Coomassie brilliant blue on PAGE: one was very clear, and the other was faint. Molecular weight for purified PPO was estimated to be about 41 000 by gel filtration. The enzyme quickly oxidized dopamine, and its Km value (Michaelis constant) for dopamine was 3.9 mM. Optimum pH was 6.5 and the PPO activity was quite stable in the range of pH 5-11 for 48 h. The enzyme had an optimum temperature at 30 degrees C and was stable up to 60 degrees C after heat treatment for 30 min. The enzyme activity was strongly inhibited by sodium diethyldithiocarbamate, potassium cyanide, L-ascorbic acid, and cysteine at 1 mM. Under a low buffer capacity, the enzyme was also strongly inhibited by citric acid and acetic acid at 10 mM.
 ...
PMID:Partial purification and characterization of polyphenol oxidase from banana (Musa sapientum L.) peel. 1131 78

The electrooxidation of a biotin pyrrole has allowed the formation of biotinylated polypyrrole films. Gravimetric measurements based on a quartz crystal microbalance demonstrate the efficient coupling of avidin, biotinylated polyphenol oxidase (PPO-B) and avidin-labeled alkaline phosphatase (AP-A) with the underlying biotinylated polymer film. The estimated mass increase corresponds to the anchoring of 1.6-1.8 equivalent layer of proteins. A step-by-step construction of bienzyme multilayers composed of PPO-B and AP-A was carried out on the electrode surface modified by the biotinylated polypyrrole film through avidin-biotin bridges. A spatially controlled distribution of the two enzymes was performed by the formation of one AP-A layer on 1, 5, and 10 PPO-B layers. The resulting bienzyme electrodes were applied to the determination of phenyl phosphate on the basis of amperometric detection of enzymically generated o-quinone at -0.2 V. Their analytical performances were analyzed in relation to the design of the enzyme architectures and in comparison with the amperometric behavior of the monoenzymatic electrodes (PPO-B electrode and AP-A electrode). It appears that at the 10-layer-PPO-B polypyrrole electrode only 4% of phenol is transformed, whereas 42-69% of phenyl phosphate is enzymatically consumed and detected at the AP-A polypyrrole electrode, depending on the enzyme activity. For the bienzymatic AP-A/PPO-B polypyrrole electrodes, the activity of each immobilized enzyme clearly affects the biosensor performance, the main limiting factor being the very low efficiency of PPO-B at pH 8.8.
 ...
PMID:Elaboration and characterization of spatially controlled assemblies of complementary polyphenol oxidase-alkaline phosphatase activities on electrodes. 1146 32

The biochemical anti-herbivore defense of trembling aspen (Populus tremuloides Michx.) was investigated in a molecular analysis of polyphenol oxidase (PPO; EC 1.10.3.2). A PPO cDNA was isolated from a trembling aspen wounded leaf cDNA library and its nucleotide sequence determined. Southern analysis indicated the presence of two PPO genes in the trembling aspen genome. Expression of PPO was found to be induced after herbivory by forest tent caterpillar, by wounding, and by methyl jasmonate treatment. Wound induction was systemic, and occurred in unwounded leaves on wounded plants. This pattern of expression is consistent with a role of this enzyme in insect defense. A search for potential PPO substrates in ethanolic aspen leaf extracts using electron spin resonance (ESR) found no pre-existing diphenolic compounds. However, following a brief delay and several additions of oxygen, an ESR signal specific for catechol was detected. The source of this catechol was most likely the aspen phenolic glycosides tremulacin or salicortin which decomposed during ESR experiments. This was subsequently confirmed in experiments using pure salicortin.
 ...
PMID:Polyphenol oxidase and herbivore defense in trembling aspen (Populus tremuloides): cDNA cloning, expression, and potential substrates. 1147 16

The amperometric response toward phenol of PPO-based rotating disk bioelectrodes is analyzed on the basis of a kinetic model taking into account internal and external mass transport effects and a CEC' electroenzymatic mechanism. Monophenolase activity of PPO catalyses the oxidation of phenol to o-quinone (step C). o-Quinone can then enter an amplification recycling process involving electrochemical reduction (step E) and enzymatic reoxidation (step C': catecholase activity). The rate-limiting steps such as monophenolase activity, catecholase recycling, permeability of the membrane, and activity and accessibility of the catalytic enzyme sites are theoretically considered and experimentally demonstrated for different electrode configurations including PPO immobilized in Laponite hydrogels and layer-by-layer self-assembled multilayers of PPO and poly(diallyldimethylammonium).
 ...
PMID:Amplification of amperometric biosensor responses by electrochemical substrate recycling. 3. Theoretical and experimental study of the phenol-polyphenol oxidase system immobilized in Laponite hydrogels and layer-by-layer self-assembled structures. 1147 17

Molecular characterisation of clonal apple rootstocks using isozymes was carried out to identify isozyme polymorphism in seven clonal apple rootstocks and to identify the most characteristic and stable enzyme markers for each individual rootstock. Five enzyme systems were studied out of which polyphenol oxidase, malate dehydrogenase, acid phosphatase and peroxidase were useful in discriminating among the rootstocks. The peroxidase enzyme system showed maximum variation and esterase showed the least variation among the rootstocks. Out of seven rootstocks, three were distinguished on the basis of one enzyme system only (M.3 with MDH or PER, M.7 with PPO or PER and MM. 111 with MDH). Out of the sixteen loci studied seven were found to be polymorphic. Genetic variation among the rootstocks was explained on the basis of various parameters. The percentage of polymorphic loci varied from 13.33 to 35.71 per cent.
 ...
PMID:Distinguishing clonal apple rootstocks by isozymes banding patterns. 1190 9

Cut tissues from distinct anatomical locations in iceberg lettuce (Lactuca sativa L.) were subjected to washing in cold (4 degrees C) and warm (47 degrees C) water with or without chlorine to assess their propensity to discoloration during storage. Total protein (Bradford method) and phenolic (TPH; Folin-Ciocalteu method) contents and polyphenol oxidase (PPO; spectrophotometric method using catechol as a substrate), peroxidase (POD; guaiacol substrate), and phenylalanine ammonia-lyase (PAL; phenylalanine substrate) activities were determined in photosynthetic and vascular tissue from outer and inner leaves. Unprocessed photosynthetic and inner leaf tissues had significantly higher (P < 0.05) levels of protein and TPH and PPO and POD activities than vascular and outer leaf tissues. PAL activities (on a fresh weight basis) were similar in all tissues. Changes in browning (light reflectance measurement) and phenolic metabolism in all four tissue types were observed during aerobic storage at 5 degrees C over 10 days. PAL activity increased in all tissues after 1-2 days of storage and then gradually decreased. POD activity also increased steadily for the storage duration. Protein content and PPO activity remained constant. Edge browning (measured with a Minolta Chroma Meter) and TPH increased in all tissues, especially in outer vascular tissue. Cut photosynthetic and vascular tissues washed at 4 and 47 degrees C with and without 100 microg mL(-1) chlorine for 3 min were analyzed during 7 days in storage at 5 degrees C. Enzyme activities and accumulation of phenolics in all tissues washed at 47 degrees C were significantly (P < 0.05) lower compared to controls or tissues washed at 4 degrees C. Chlorine had no additional effect at 47 degrees C but significantly (P < 0.05) reduced browning and accumulation of phenolics in lettuce washed at 4 degrees C. These results showed that inherent differences between tissues affect phenolic metabolism and browning in stored, fresh-cut lettuce.
 ...
PMID:Effect of wash water temperature and chlorination on phenolic metabolism and browning of stored iceberg lettuce photosynthetic and vascular tissues. 1213 68


1 2 3 4 5 6 7 8 9 Next >>