IQUIFIB   02644
INSTITUTO DE QUIMICA Y FISICOQUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Differential Exposure of Hydrophobic Domains After Calmodulin and Phosphatidic Acid Activation
Autor/es:
JUAN PABLO F. ROSSI, IRENE C. MANGIALAVORI, ANA MARÍA VILLAMIL GIRALDO, MARÍA F. PIGNATARO, MARIELA S. FERREIRA-GOMES, ARIEL J. CARIDE
Lugar:
Baltimore
Reunión:
Congreso; 55th Annual Meeting of Biophysical Society. Baltimore, Maryland, 5-9, Marzo, 2011; 2011
Institución organizadora:
Biophysical Society
Resumen:
PMCA Differential
Exposure of Hydrophobic Domains After Calmodulin and Phosphatidic Acid
Activation
Juan Pablo F. Rossi,
Irene C. Mangialavori, Ana María Villamil Giraldo, María F. Pignataro, Mariela S. Ferreira-Gomes, Ariel J.
Caride
The exposure of plasma membrane calcium pump
(PMCA) to surrounding phospholipids was assessed by measuring the incorporation of the photoactivatable
phosphatidylcholine analogue [125I]TID-PC/16 to the protein. In the
presence of Ca2+ both calmodulin (CaM) and phosphatidic acid (PA)
greatly decreased the incorporation of [125I]TID-PC/16 to PMCA.
Proteolysis of PMCA with V8 protease results in 3 main fragments: N which includes transmembrane segments
M1 and M2, M which includes M3 and M4
and C which includes M5 to M10.
CaM decreased the level of incorporation of [125I]TID-PC/16
to fragments M and C, while phosphatidic acid decreased the
incorporation of [125I]TID-PC/16 to fragments N and M. This suggests
that the conformational changes induced by binding of CaM or PA extend to the
adjacent transmembrane domains. Interestingly, this result also denotes
differences between the active conformations produced by CaM and PA. To verify
this point, we measured RET between PMCA labeled with eosin-isothiocyanate at the
ATP-binding site and the phospholipid Rho-PE included in PMCA micelles. CaM
decreased the efficiency of the energy transfer between these two probes while
PA did not. This result indicates that activation by CaM increases the distance
between the ATP-binding site and the membrane, but PA does not affect this
distance. Our results disclose main differences between PMCA conformations
induced by CaM or PA and show that those differences involve transmembrane
regions. With grants of CONICET, ANPCYT and UBACYT