Purification and Properties of Plasma Membrane (Ca2 +-Mg2 + ) -ATPasefrom Synaptosomes of Pig Brain
- VernacularTitle:猪脑突触体质膜(Ca2+-Mg2+)-ATPase的分离纯化与性质
- Author:
Xiaoxuan FAN
;
Xujia ZHANG
- Publication Type:Journal Article
- From:
Progress in Biochemistry and Biophysics
2001;28(1):90-93
- CountryChina
- Language:Chinese
-
Abstract:
Synaptosomes were isolated from pig brain by homogenization, differential centrifugation and sucrose gradient centrifugation. After synaptosome lysis in hypoosmotic buffer, the plasma membrane vesicles were collected. Following the solubilization of plasma membrane vesicles in Triton X-100, the solubilized protein was applied to calmodulin affinity chromatography colurnn, and the delipidated plasma membrane Ca2 + -ATPase was purified to nearly homogeneity. The novel feature of this purification is the use of large affinity column and heavy washing to facilitate the purified Ca2+ -ATPase with higher activity and protein yield. The specific activity of the purified Ca2+ -ATPase was recovered to a maximum of 3.32 μmol· mg-1· min-1 after incubation with asolectin. Silver staining of SDS-PAGE revealed a single protein band around Mr 140 000, showing the purity was over 90 %. Different Ca2 + concentrations dramatically affect the specific activity of Ca2 + -ATPase.
