Purification of the Plasma Membrane Ca2+-ATPase from Radish Seedlings by Calmodulin-Agarose Affinity Chromatography
Articolo
Data di Pubblicazione:
1998
Citazione:
Purification of the Plasma Membrane Ca2+-ATPase from Radish Seedlings by Calmodulin-Agarose Affinity Chromatography / C. Bonza, A. Carnelli, M.I. De Michelis, F. Rasi-Caldogno. - In: PLANT PHYSIOLOGY. - ISSN 0032-0889. - 116:2(1998), pp. 845-851. [10.1104/pp.116.2.845]
Abstract:
The Ca2+-ATPase of the plasma membrane (PM) of germinating radish (Raphanus sativus I.) seeds was purified by calmodulin (CaM)-affinity chromatography using a batch procedure. PM purified by aqueous two-phase partitioning was solubilized with n-dodecyl β-D-maltoside and applied to a CaM-agarose matrix. After various washings with decreasing Ca2+ concentrations, the Ca2+-ATPase was eluted with 5 mM ethylenediaminetetraacetate (EDTA). The EDTA-eluted fraction contained about 25% of the loaded Ca2+-ATPase activity, with a specific activity 70-fold higher than that of the starting PM fraction. The EDTA-eluted fraction was highly enriched in a 133-kD polypeptide, which was identified as the PM Ca2+-ATPase by 125I-CaM overlay and fluorescein-isothiocyanate labeling. The PM Ca2+-ATPase cross-reacted with an antiserum against a putative Ca2+-ATPase of the Arabidopsis thaliana chloroplast envelope.
Tipologia IRIS:
01 - Articolo su periodico
Keywords:
Calcium-pumping atpases; beet beta-vulgaris; red beet; functional reconstitution; signal-transduction; brassica-oleracea; stimulated atpase; CA2+ transport; adapted cells; messenger-RNA
Elenco autori:
C. Bonza, A. Carnelli, M.I. De Michelis, F. Rasi-Caldogno
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