Studies on alkaline phosphatase. Phosphorylation of calf intestinal alkaline phosphatase by 32P-labelled pyrophosphate

Abstract

1. A purified preparation of alkaline phosphatase from calf-intestinal mucosa was phosphorylated by ³²P-labelled PP_i at a serine residue on the enzyme. Under the conditions employed, up to 0·15μm-labelled sites were obtained from 1μm-[³²P]PP_i. 2. The phosphorylated enzyme was labile, the rate of dephosphorylation being similar to the overall rate of substrate hydrolysis. 3. A stopped-flow technique was used to determine the number of phosphomonoesterase active sites, which agreed with the number of ³²P-labelled sites. 4. It is concluded that calf-intestinal alkaline phosphatase is both a phosphomonoesterase and a pyrophosphatase.