Purification and Properties of Alkaline Phosphatase with Protein Phosphatase Activity from Saccharomyces cerevisiae

An alkaline phosphatase (A LPase) from Saccharomyces cerevisiae strain 257 was purified 345-fold with specific activity of 54 533 nmol x min“ 1 x mg protein-1 . It was shown to be a dimeric protein (apparent mol. wt. approx. 130 kDa) with optimum activity at pH 8.6 8.8 and good stability at 50 °C. The A LPase was a non-specific enzyme hydrolyzing a wide vari­ ety of monophosphate esters. The enzyme showed protein phosphatase activity and this activ­ ity was not Mg2+ dependent in contrast to /?-nitrophenyl phosphate (pNPP) activity. The Km value for pNNP hydrolysis was determined to be 2.2 x 10“ 5 m. Orthophosphate inhibited the enzyme in a competitive mode with the of 2.3 x 10“4 m. Phosphate transfer of the A LPase is almost zero with all alcohols tested except for Tris.