RT Journal Article SR Electronic T1 Purification of the Plasma Membrane Ca2+-ATPase from Radish Seedlings by Calmodulin-Agarose Affinity Chromatography JF Plant Physiology JO Plant Physiol. FD American Society of Plant Biologists SP 845 OP 851 DO 10.1104/pp.116.2.845 VO 116 IS 2 A1 Bonza, Cristina A1 Carnelli, Antonella A1 De Michelis, Maria Ida A1 Rasi-Caldogno, Franca YR 1998 UL http://www.plantphysiol.org/content/116/2/845.abstract AB The Ca2+-ATPase of the plasma membrane (PM) of germinating radish (Raphanus sativus L.) seeds was purified by calmodulin (CaM)-affinity chromatography using a batch procedure. PM purified by aqueous two-phase partitioning was solubilized withn-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 thalianachloroplast envelope. Brij 58polyoxyethylene-20-cetyl etherCaMcalmodulinFITCfluorescein isothiocyanatePMplasma membrane