Abstract
Candida utilis alkaline phosphatase has been detected in vacuoles. Liberation of the vacuoles was carried out by protoplast disruption under isotonic conditions. The polybase DEAE-dextran was used to induce damage to the yeast plasmalemma. The vacuoles were purified by centrifugation on sorbitol-Ficoll gradients. Alkaline phosphatase from a purified fraction of vacuoles was characterized after gel filtration on Sephadex G-200. We have found 15 mU of enzyme activity per 108 vacuoles. This enzyme activity elutes on Sephadex G-200 at a volume-to-void-volume ratio of 1.65. The approximate molecular weight is 1.35×105. TheK m value forp-nitrophenyl-phosphate is 2.5×10−3 M. The pH for maximum activity is 8.9, and the enzyme is stable at pH values between 7.0 and 9.0. Rapid inactivation occurs at temperatures above 45°C. The enzyme catalyzes the hydrolysis of phosphomonoester bonds of a wide variety of molecules, especially polyphosphates. Thus, vacuolar polyphosphates are probably the natural substrate of this enzyme. Orthophosphate, arsenate, ethylenediaminetetraacetate, molybdate, and borate act as inhibitors. Fluoride is not an inhibitor, and the activity is not affected byp-hydroxymercuribenzoate. Some metal ions also affect the activity of vacuolar alkaline phosphatase. This may indicate that this enzyme is a metalloprotein.
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Fernandez, M.P., Gascon, S. & Schwencke, J. Some enzymatic properties of vacuolar alkaline phosphatase from yeast. Current Microbiology 6, 121–126 (1981). https://doi.org/10.1007/BF01569016
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DOI: https://doi.org/10.1007/BF01569016