The purification and molecular characterization of alkaline phosphatases from chondrocytes and matrix vesicles of bovine fetal epiphyseal cartilage

摘要: Abstract Intact matrix vesicles and chondrocytes were obtained from a crude coilagenase digest of bovine fetal epiphyseal cartilage. Alkaline phosphatase activity the was purified 1800-fold to homogeneity. This accomplished by solubilizing in sodium deoxycholate, followed subsequent gel filtration on Sepharose 6B DE-52 cellulose chromatography. A 30-fold purification vesicle alkaline solubilization with extraction n-butanol 6B. The chondrocyte possessed specific 900 μmoles p-nitrophenyl phosphate hydrolyzed/minute/mg protein at pH 10.5, while has 300 p-nitrophenyi enzyme. native molecular weight chondrocytic determined be 141,000 310,00, respectively, sucrose density gradient centrifugation. functional subunit phosphatase, identical isolated could derived intact enzyme chromatography presence 1 mM β-mercaptoethanol SDS polyacrylamide electrophoresis demonstrated that had one band approximately 18,000 two bands, other 45,000 weight.