THE METAL ION ACTIVATION OF THE ALKALINE BETA-GLYCEROPHOSPHATASE OF RABBIT SMALL INTESTINE.

1. A fraction of intestinal epithelial cells from rabbit small intestine that contained nuclei and microvillus membranes served as a source of alkaline-beta-glycerophosphatase activity. 2. The greater part of the enzyme activity could be released from the subcellular particles by disintegration of the latter, followed by centrifugation at 40000g and butanol extraction of the resulting sediment. 3. Further purification of the enzyme was achieved by diethylaminoethylcellulose chromatography and by gel filtration. 4. Dialysis of the purified enzyme preparations against EDTA gave an essentially inactive enzyme. High activity could be restored by adding Zn(2+)+Mg(2+), Zn(2+)+Co(2+), Mg(2+)+Co(2+) or Co(2+) alone to these inactive preparations. Neither Zn(2+) nor Mg(2+) added singly to the assay system restored more than a small part of the enzyme activity. 5. The optimum Zn(2+) concentration was about 0.2-1m-equiv./l., whereas Mg(2+) and Co(2+) had optimum concentrations about 30-60m-equiv./l. 6. If added in excess of the optimum concentration, Zn(2+) strongly inhibited the enzyme under all conditions tested. 7. In the presence of an optimum concentration of Co(2+) (33m-equiv./l.) in tris buffer at the optimum pH (8.8 at 37 degrees ), K(m) for the beta-glycerophosphatase was 0.3mm.