Purification and subunit structure of alkaline phosphatase from bovine enamel organ.

A highly-purified alkaline-phosphatase preparation having a specific activity of 703 U/mg protein was obtained from bovine enamel organ by a series of procedures: butanol extraction, isoelectric and acetone precipitation, ion-exchange, concanavalin A affinity and gel-filtration chromatography. The purified enzyme showed the same properties as kidney-type isozyme and contained carbohydrate moieties which react with concanavalin A. Analysis by polyacrylamide gel electrophoresis showed that the purified enzyme split into half the size of the native molecule (160,000 in mol. wt) after being heated in sodium dodecyl sulphate (SDS) solution and the subunit had no catalytic activity. The results indicate that the enzyme is a dimeric glycoprotein comprised of two identical subunits, each having a molecular weight of 80,000.