THE EFFECT OF ACIDIC LIPIDS ON THE Ca2+-INDEPENDENT PHOSPHATASE ACTIVITY OF THE PLASMA MEMBRANE Ca2+ PUMP.

MAZZITELLI LUCIANA ROMINA; ADAMO HUGO PEDRO

Pinamar, Pcia. Bs.As

Congreso; XLI Reunion Anual de la Sociedad Argentina de Investigacion en Bioquimica y Biologia Molecular (SAIB); 2005

Sociedad Argentina de Investigacion en Bioquimica y Biologia Molecular (SAIB)

The effect of acidic lipids on the Ca2+-independent phosphatase activity of the plasma membrane Ca2+ pump. Luciana R. Mazzitelli and Hugo P. Adamo, IQUIFIB-Facultad de Farmacia y Bioquímica (UBA-CONICET), Buenos Aires. lrmazzitelli@hotmail.com The purified plasma membrane Ca2+ pump PMCA reconstituted in phosphatidylcoline (PC) liposomes was able to hydrolyze p-nitrophenylphosphate (pNPPase) at a rate of 0.2 µmol/mg/min in the absence of Ca2+, ATP and calmodulin. Reconstitution in mixture of acidic lipids (BE) increased the pNPPase 25 fold. Ca2+ inhibited the PC-pNPPase following a biphasic curve with KCa1= 0.52 µM and KCa2= 300 µM. In contrast the inhibition of the BE-pNPPase by Ca2+ exhibited only the high affinity component.  The lipidic composition of the liposomes did not change the apparent affinity for pNPP. However Mg2+ activated the BE-pNPPase with much higher aparent affinity than the PC-pNPPase  (KMg 0.59 µM and 8.44 µM respectively).  The results are consistent with the idea that the E2 form (absence Ca2+) of the PMCA catalyses the pNPP hydrolysis. Ca2+ binding to the transport site inhibit this activity.  In the absence of  Ca2+ and acidic lipids the PMCA would reside in an E2 inhibited state with low pNPPase activity and low affinity for Mg2+.  Under these conditions only a portion of the enzyme would bind Ca2+ with high affinity.  [Supported by grants of CONICET and UBA].