Ascorbate Oxidase (ascorbate + oxidase)

Distribution by Scientific Domains


Selected Abstracts


Physico-chemical properties of molten dimer ascorbate oxidase

FEBS JOURNAL, Issue 22 2006
Eleonora Nicolai
The possible presence of dimeric unfolding intermediates might offer a clue to understanding the relationship between tertiary and quaternary structure formation in dimers. Ascorbate oxidase is a large dimeric enzyme that displays such an intermediate along its unfolding pathway. In this study the combined effect of high pressure and denaturing agents gave new insight on this intermediate and on the mechanism of its formation. The transition from native dimer to the dimeric intermediate is characterized by the release of copper ions forming the tri-nuclear copper center located at the interface between domain 2 and 3 of each subunit. This transition, which is pH-dependent, is accompanied by a decrease in volume, probably associated to electrostriction due to the loosening of intra-subunit electrostatic interactions. The dimeric species is present even at 3 × 108 Pa, providing evidence that mechanically or chemically induced unfolding lead to a similar intermediate state. Instead, dissociation occurs with an extremely large and negative volume change (,V , ,200 mL·mol,1) by pressurization in the presence of moderate amounts of denaturant. This volume change can be ascribed to the elimination of voids at the subunit interface. Furthermore, the combination of guanidine and high pressure uncovers the presence of a marginally stable (,G , 2 kcal·mol,1) monomeric species (which was not observed in previous equilibrium unfolding measurements) that might be populated in the early folding steps of ascorbate oxidase. These findings provide new aspects of the protein folding pathway, further supporting the important role of quaternary interactions in the folding strategy of large dimeric enzymes. [source]


On-line biosensors for simultaneous determination of glucose, choline, and glutamate integrated with a microseparation system

ELECTROPHORESIS, Issue 18 2003
Guoyue Shi
Abstract An effective microseparation system integrated with ring-disc electrodes and two microfluidic devices was fabricated for in vivo determination using a microdialysis pump. The major interference of ascorbic acid (AA) was excluded by direct oxidation with ascorbate oxidase. Glucose, glutamate, and choline were successfully determined simultaneously through the biosensors modified with a bilayer of osmium-poly(4-vinylpyridine)gel-horseradish peroxidase (Os-gel-HRP)/glucose oxidase (GOD), glutamate oxidase (GlutaOD) or choline oxidase (ChOD). To stabilize the biosensors, 0.2% polyethylenimine (PEI) was mixed with the oxidases. The cathodic currents of glucose, glutamate, and choline biosensors started to increase after the standard solutions were injected into the microseparation system. The on-line biosensors show a wide calibration range (10,7,10,5 mol/L) with a detection limit of 10,8 mol/L at the working potential of ,50 mV. The variations of glucose, glutamate, and choline were determined simultaneously in a free moving rat when we perfused the medial frontal cortex with 100 ,mol/L N -methyl- D -aspartate (NMDA) solution, which is the agonist of the NMDA receptor. [source]


Physico-chemical properties of molten dimer ascorbate oxidase

FEBS JOURNAL, Issue 22 2006
Eleonora Nicolai
The possible presence of dimeric unfolding intermediates might offer a clue to understanding the relationship between tertiary and quaternary structure formation in dimers. Ascorbate oxidase is a large dimeric enzyme that displays such an intermediate along its unfolding pathway. In this study the combined effect of high pressure and denaturing agents gave new insight on this intermediate and on the mechanism of its formation. The transition from native dimer to the dimeric intermediate is characterized by the release of copper ions forming the tri-nuclear copper center located at the interface between domain 2 and 3 of each subunit. This transition, which is pH-dependent, is accompanied by a decrease in volume, probably associated to electrostriction due to the loosening of intra-subunit electrostatic interactions. The dimeric species is present even at 3 × 108 Pa, providing evidence that mechanically or chemically induced unfolding lead to a similar intermediate state. Instead, dissociation occurs with an extremely large and negative volume change (,V , ,200 mL·mol,1) by pressurization in the presence of moderate amounts of denaturant. This volume change can be ascribed to the elimination of voids at the subunit interface. Furthermore, the combination of guanidine and high pressure uncovers the presence of a marginally stable (,G , 2 kcal·mol,1) monomeric species (which was not observed in previous equilibrium unfolding measurements) that might be populated in the early folding steps of ascorbate oxidase. These findings provide new aspects of the protein folding pathway, further supporting the important role of quaternary interactions in the folding strategy of large dimeric enzymes. [source]


Degradative enzymatic activities in fresh-cut blood-orange slices during chilled-storage

INTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 5 2009
Anna Eghle Catalano
Summary Blood-orange fruits are suitable to fresh-cut fruit production because of their chemical compositions. Nevertheless, the main limitation of using freshly cut oranges is their susceptibility to juiciness loss and ascorbic acid degradation because of enzymatic alterations. The aim of this work is: to identify some of the enzymes causing the qualitative decay in blood-orange slices during 15 days of chilled storage (at 4 ± 0.5 °C and 85% RH); to investigate the susceptibility to the previous alterations of five blood-orange clones (Moro nucellare, Sanguinello nucellare, Tarocco arcimusa, Tarocco gallo and Tarocco meli) to select the most suitable one for fresh-cut production. The enzymes studied were: pectinmethylesterase (PME) as index of juiciness loss, ascorbate oxidase (AAO) as index of ascorbic acid's degradation and polyphenol oxidase (PPO) as browning index. As far as we know, the changes of AAO activity during chilled storage of blood-orange fresh-cut slices has not previously reported and studied. Different clones showed different enzymatic activities and quality changes during chilled-storage. In particular a low juiciness loss in orange slices was correlated with a lower PME activity, as described in T. meli clone, while a high degradation of ascorbic acid was correlated with an higher AAO activity, as described in T. gallo clone; PPO activity seemed to have no significant action in quality degradation. Tarocco meli was the most suitable clone to the fresh-cut blood-orange production because it has the lowest enzymatic activity (PME, PPO and AAO) and the highest sensorial quality. [source]


The oxidation of l -ascorbic acid catalysed by pear tyrosinase

PHYSIOLOGIA PLANTARUM, Issue 1 2000
Juan Carlos Espín
The ability of partially purified pear tyrosinase (PPO) to catalyse the oxidation of l -ascorbic acid (AA) has been reported here for the first time. The ascorbate oxidase activity of PPO was studied by oxymetric assays. The activity was linearly related to the enzyme concentration with a Michaelis constant (Km) for AA of 0.55±0.03 mM at pH 7. The stoichiometry was found to be 1:2 (O2:AA). The action of the PPO inhibitors tropolone and sodium chloride was studied to exclude a possible interference of endogenous pear ascorbate oxidase in the oxidation of AA. A possible role of the ,AA/PPO' system in the browning of pears is proposed. [source]