Alkaline phosphatase of Physarum polycephalum is insoluble.
The plasmodia of Physarum polycephalum grow as multinucleated cells in the presence of sufficient humidity and nutriment. Under non-illuminating conditions, stresses such as low temperature or high concentrations of salts transform the plasmodia into spherules whereas dehydration induces sclerotization. Some phosphatases including protein phosphatase and acid phosphatase have been purified from the plasmodia, but alkaline phosphatase remains to be elucidated. Phosphatase of the plasmodia, spherules and sclerotia was visualized by electrophoresis gel-staining assay using 5-bromo-4-chloro-3-indolyl phosphate. Insoluble fractions of the sclerotia were abundant in phosphatase activity. The phosphatase which was extracted by nonionic detergent was subjected to column chromatography and preparative electrophoresis. Purified phosphatase showed the highest activity at pH 8.8, indicating that this enzyme belongs to alkaline phosphatase. The apparent molecular mass from sodium dodecyl sulfate-polyacrylamide gel electrophoresis under non-reducing condition was estimated to be 100 kDa whereas that under reducing was 105 kDa. An amount of 1% sodium dodecyl sulfate or 0.5 M NaCl had no effects on the activity although the phosphatase showed heat instability, Mg(2+)-dependency and sensitivity to 2-glycerophosphate or NaF. The extracting conditions and enzymatic properties suggest that this alkaline phosphatase which is in a membrane-bound form plays important roles in phosphate metabolism.
Furuhashi
Kiyoshi
K
Faculty of Education, Shiga University, Hiratsu 2-5-1, Otsu, Shiga, 520-0862, Japan. furuhasi@sue.shiga-u.ac.jp
D000469:Alkaline Phosphatase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002845:Chromatography; D003067:Coenzymes / Q000494:pharmacology; D004586:Electrophoresis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D005994:Glycerophosphates / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D015636:Magnesium Chloride / Q000494:pharmacology; D008970:Molecular Weight; D016839:Physarum polycephalum / Q000201:enzymology*; D015800:Protozoan Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D012969:Sodium Fluoride / Q000494:pharmacology; D012995:Solubility; D013696:Temperature
D003067:Coenzymes; D004791:Enzyme Inhibitors; D005994:Glycerophosphates; D015800:Protozoan Proteins; D015636:Magnesium Chloride; D012967:Sodium Dodecyl Sulfate; D012965:Sodium Chloride; D012969:Sodium Fluoride; D000469:Alkaline Phosphatase