Choline Oxidase from Alcaligenes sp.

Isoelectric point:4.1 ± 0.1
Michaelis constants:2.84 x 10¯³M (Choline), 5.33 x 10¯³M (Betaine aldehyde)
Structure:One mol of FAD is covalently bound to mol of the enzyme
Inhibitors:p-Chloromercuribenzoate, Cu⁺⁺, Co⁺⁺, Hg⁺⁺, Ag⁺
Optimum pH:8.0 – 8.5
Optimum temp:40 – 45°C
pH Stability:pH 7.0 – 9.0 (30°C, 2hr)
Thermal stability:Below 37°C (pH 7.5, 10min)

Choline oxidase from Alcaligenes sp. has been used:

• in a study to investigate a choline biosensor constructed with a chitinous membrane and its application in measuring cholinesterase inhibitory activities
• has also been used in spectroscopic studies on the photoreaction of choline oxidase with covalently bound flavin.
• as a component of phosphate buffer saline (PBS) buffer for measurement of choline acetyltransferase (ChAT) activity in brain tissue homogenates
• to measure the choline generated by autotaxin (ATX) by ATX activity assay and phospholipase-D (PLD) by PLD activity assay

Choline oxidase catalyzes the four-electron oxidation of choline to glycine-betaine, with betaine-aldehyde as the intermediate and molecular oxygen as th eprimary electron acceptor. The enzyme can also accept betaine-aldehyde as a substrate. This allows the study of the reaction mechanism for the conversion of choline to the aldehyde intermediate, & of betaine-aldehyde to glycine-betaine. The enzyme is a flavoprotein with a molecular weight of approx. 72,000 Da (gel filtration) or 66,000 Da (SDS gel electrophoresis).

One unit will form 1 μmole of H₂O₂ with oxidation of 1 μmole of choline to betaine aldehyde per min at pH 8.0 at 37 °C. Note: During the conversion of choline to betaine by choline oxidase, 2 μmoles of H₂O₂ are produced for every μmole of choline.