H+/Ca2+ exchange driven by the plasma membrane Ca2+‐ATPase of Arabidopsis thaliana reconstituted in proteoliposomes after calmodulin‐affinity purification

FEBS Letters

Bonza, Maria Cristina¹  Luoni, Laura¹  De Michelis, Maria Ida¹ 

[1] Dipartimento di Biologia ‘L. Gorini’, Università di Milano, Centro di Studio del CNR per la Biologia Cellulare e Molecolare delle Piante, via G. Celoria 26, 20133 Milan, Italy

关键词: Ca2+-ATPase;    Calmodulin;    H+/Ca2+ antiport;    Plasma membrane;    Proteoliposome;    Pyranine;    Arabidopsis thaliana;    BTP;    bis-tris propane (1;    3-bis[tris (hydroxymethyl)methylamino]-propane);    CaM;    calmodulin;    EO;    eosin Y;    FCCP;    carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone;    PM;    plasma membrane;   

DOI  :  10.1016/S0014-5793(00)02065-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The plasma membrane Ca²⁺-ATPase was purified from Arabidopsis thaliana cultured cells by calmodulin (CaM)-affinity chromatography and reconstituted in proteoliposomes by the freeze-thaw sonication procedure. The reconstituted enzyme catalyzed CaM-stimulated ⁴⁵Ca²⁺ accumulation and H⁺ ejection, monitored by the increase of fluorescence of the pH probe pyranine entrapped in the liposomal lumen during reconstitution. Proton ejection was immediately reversed by the protonophore FCCP, indicating that it is not electrically coupled to Ca²⁺ uptake, but it is a primary event linked to Ca²⁺ uptake in the form of countertransport.

【 授权许可】

Unknown