In vitro Ca deposition by rat matrix vesicles: Is the membrane association of alkaline phosphatase essential for matrix vesicle-mediated calcium deposition?

Abstract

1.1. Phosphatidylinositol phospholipase C (PI-PLC) treatment of rachitic rat matrix vesicles (MVs) released about 80% of membrane-bound alkaline phosphatase (ALP), AMPase, PPiase into the media.2.2. About 20% hydrolytic activity was not released from MV membranes by PI-PLC treatment.3.3. SDS-polyacrylamide gel electrophoresis and Western blot analysis showed only one immunoreactive protein corresponding to the molecular weight of ALP present in the soluble fraction after PI-PLC treatment.4.4. The specific activity of the released ALP was at least 5-fold higher than the residual activity.5.5. After PI-PLC treatment, MVs also demonstrated an 80% reduction of AMP- or βGP-dependent calcium deposition.6.6. The soluble fraction containing 80% of ALP activity was unable to support calcium deposition. The mixing of the soluble and insoluble fractions after PI-PLC treatment failed to fully restore calcium-depositing activity.