AAO

Ascorbate oxidase (ascorbic acid oxidase) is a copper-containing enzyme located in the cytoplasm or bound to the cell wall. Ascorbic acid oxidase is coupled with other redox reactions to function as a terminal oxidase, which catalyzes the oxidation of ascorbic acid, has an anti-aging effect, and plays an important role in the metabolism of substances in plants. Pyruvic acid, isocitric acid, α-ketoglutaric acid, malic acid, glucose-6-phosphate, and 6-phosphogluconate can all remove H protons by dehydrogenase and transfer H protons to coenzyme. Then, glutathione is used to transfer the H proton to ascorbic acid. Under the action of ascorbate oxidase, ascorbic acid is oxidized to O₂, which combines with H protons to form water.

Structure

Ascorbate oxidase is a homodimeric enzyme with a molecular mass of 70 kDa and 552 amino acid residues per subunit (zucchini). The folding of all three domains is of a similar β-barrel type. The mononuclear copper site is located in domain 3 and the trinuclear copper species is bound between domains 1 and 3. A binding pocket for the reducing substrate that is complementary to an ascorbate molecule is located near the type-1 copper site and accessible from solvent. A broad channel providing access from solvent to the trinuclear copper species, which is the binding and reaction site for the dioxygen is present in ascorbate oxidase.

Enzyme Activity Assay

A commonly used method for determining ascorbate oxidase activity is the iodometric method, which is a classical method with a simple instrument and high accuracy. The second method is spectrophotometry, which has high sensitivity and accuracy, good reproducibility, wide application and good selectivity. The third method is the oxygen electrode analysis method, which has high sensitivity and good accuracy, but the oxygen electrode is very sensitive to temperature changes, and it is necessary to maintain a constant temperature during the measurement. The fourth method is pulse polarography, which is simple in operation and reproducible, consistent with the results of the iodometric method.

Influence Factors

Applications

Ascorbate oxidase is of great significance in plant physiology. Studies have shown that ascorbate oxidase activity is associated with wheat disease resistance and pepper resistance to aphids. After different treatments, some fruits such as kiwi, thorn pear, apple, and winter jujube can be kept fresh for a long time. If the metal ion stress conditions such as aluminum ion and calcium ion are used, the enzyme activity can be improved to some extent; in the presence of small organic molecules, the activity also changes to different degrees, which provides a reference for planting under different cultivation conditions.

Reference