Active Transport of Divalent Cations by the Plasma Membrane Calcium Pump

MALLKU ONTIVEROS ; IRENE MANGIALAVORI; MARIELA FERREIRA -GOMES; JUAN PABLO ROSSI

San Miguel-Tucuman

Congreso; XLI. Reunión Anual de la Sociedad Argentina de Biofísica (SAB); 2012

Sociedad de Biofísica Argentina

The plasma membrane calcium pump (PMCA) is widely distributed along the eukaryotic cells. This pump transports actively Ca2+ from cytoplasm to extracellular medium coupled to the hydrolysis of ATP maintaininga low intracellular concentration of the cation. PMCA is regulated by theCa2+-calmodulin complex(Ca2+-CaM). In the absence of CaM the pump is auto-inhibited; while binding of CaM to the C-terminal domain produces the activation ofthe pump1. The aim of this work is to study the transport of different divalent cations by PMCA for both the auto-inhibited and the activated state. Purified PMCA preparations and inside-out vesicles (IOVS) were obtained from human erythrocytesmembranes. We measured the Cation-ATPaseactivity as function of the ions Ca2+, Sr2+, Ba2+ and Pb2+, in the presence and the absenceof CaM as well as, with the enzymepreviously activated by removing the C-terminal domain with chymotrypsin2. Additionally,we study the active transport of divalent cations into IOVS by measuring light scattering changes of the vesicles. Results shows that: (a) PMCA is able to transport divalent cations other than Ca2+; (b) These cations (C2+) are transported with different affinitiesand velocities; (c) Ba2+ are unable to bind to CaM but Ba2+-ATPase increases its activity when the auto-inhibited domain was removed. (d) Both C2+-ATPase and transport of C2+ measured by light scattering show a similar behavior revealing that both processes are coupled. These results suggest that the mechanism of expulsion ofCa2+ by PMCA would also eliminate other divalent cations. This fact indicates that PMCA could contribute to detoxification process under the eventual access of other divalent cationsinto the cell. [1] Filomatori, C. V. and Rega, A. F. (2003) J Biol Chem. 278, 22265-22271 [2] Enyedi, A., Flura, M., Sarkadi, B., Gardos, G., Carafoli, E. (1987) J. Biol. Chem. 262,6425-6430 With grants of ANPCYT, CONICET y UBACYT