Alkaline phosphatase from human liver was purified to homogeneity. The purification procedure included solubilization with butanol, fractionation with acetone, and chromatography on concanavalin A-Sepharose, DEAE-cellulose, Sephadex G-200 and DEAE-Sephadex. Purity was established by standard and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The isoelectric point of the protein was determined to be 4.0. Sephadex-gel filtration gave a mol.wt. of 146000, although a higher value was obtained in the presence of 100mM-NaC1. The subunit mol.wt. 76700, was determined by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Neuraminidase treatment resulted in two enzyme-activity bands on isoelectric-focused gels with isoelectric points of 6.6 and 6.8. The desialylated enzyme gave only one protein band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis with a subunit molecular weight indistinguishable from that of the non-neuraminidase-treated protein. The desialylated enzyme was more readily denatured by sodium dodecyl sulphate in the presence of mercaptoethanol than was the native enzyme.
Skip Nav Destination
Research Article| June 01 1976
Affinity purification and some molecular properties of human liver alkaline phosphatase
J M Trépanier ;
L E Seargeant ;
Biochem J (1976) 155 (3): 653–660.
- Views Icon Views
- Share Icon Share
J M Trépanier, L E Seargeant, R A Stinson; Affinity purification and some molecular properties of human liver alkaline phosphatase. Biochem J 1 June 1976; 155 (3): 653–660. doi: https://doi.org/10.1042/bj1550653
Download citation file:
Don't already have an account? Register
Get Access To This Article
Buy This Article