Choline oxidase was purified from the cells of Arthrobacter globiformis by fractionations with acetone and ammonium sulfate, and column chromatographies on DEAE-cellulose and on Sephadex G-200. The purified enzyme preparation appeared homogeneous on disc gel electrophoresis. The enzyme was a fiavoprotein having a molecular weight of approx. 83, 000 (gel filtration) or approx. 71, 000 (sodium dodecyl sulfate-polyacrylamide disc gel electrophoresis) and an isoelectric point (pI) around pH 4.5. Identification of the reaction products showed that the enzyme catalyzed the following reactions: choline+O₂→betaine aldehyde+H₂O₂, betaine aldehyde+O₂+H₂O→betaine+H₂O₂.
The enzyme was highly specific for choline and betaine aldehyde (relative reaction velocities: choline, 100%; betaine aldehyde, 46%; N, N-dimethylaminoethanol, 5.2%; triethanolamine, 2.6%; diethanolamine, 0.8%; monoethanolamine, N-methylaminoethanol, methanol, ethanol, propanol, formaldehyde, acetaldehyde, and propionaldehyde, 0%), Its K_m values were 1.2mM for choline and 8.7mM for betaine aldehyde. The optimum pH for the enzymic reaction was around pH 7.5.