Properties of an alkaline phosphatase from Sinclair swine melanoma.
In Sinclair swine, there is an increase in alkaline phosphatase activity in spontaneously arising melanoma tumors when compared to normal skin. While alkaline phosphatase activity could be detected in melanomas from animals 1 day old, the maximum levels of alkaline phosphatase activity occurred in tumors from animals greater than 30 days old. The alkaline phosphatase was purified from cutaneous melanomas using chloroform precipitation, Phenyl-Sepharose chromatography, and concanavalin A Sepharose chromatography approximately 146-fold, with an overall recovery of 15%. The purified enzyme exhibited optimal activity over the pH range of 8.9-10.6. The apparent Km of the enzyme for p-nitrophenyl phosphate was 0.15 mM. The enzyme exhibited a relative mobility of 0.04 in nondenaturing polyacrylamide gels. The molecular weight of the enzyme was estimated by gel filtration chromatography to be 122,000 and it was composed of two identical subunits each having a molecular weight of 67,000. The enzyme was thermolabile at 56 degrees C (T50, 18 min) and its activity was inhibited by L-homoarginine, levamisole, and vanadate, but not by L-phenylalanine or L-phenylalanylglycylglycine. These characteristics distinguished the enzyme from the intestinal isoenzyme that is found in normal swine skin but were similar to those exhibited by the porcine placental isoenzyme of alkaline phosphatase. These results suggest that the development of malignant melanoma in Sinclair swine is accompanied by the expression of a placental-like alkaline phosphatase activity.
D000469:Alkaline Phosphatase / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D000831:Animals, Newborn; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D008545:Melanoma / Q000201:enzymology / Q000662:veterinary*; D008970:Molecular Weight; D012867:Skin / Q000201:enzymology; D013552:Swine; D013553:Swine Diseases / Q000201:enzymology*
D000469:Alkaline Phosphatase