INVESTIGADORES
MANGIALAVORI Irene Cecilia
congresos y reuniones científicas
Título:
Conformational changes by ATP binding in the plasma membrane calcium pump
Autor/es:
NICOLÁS SAFFIOTI; MARIELA FERREIRA-GOMES; JUAN PABLO ROSSI; IRENE MANGIALAVORI
Lugar:
San Miguel-Tucuman
Reunión:
Congreso; XLI. Reunión Anual de la Sociedad Argentina de Biofísica (SAB); 2012
Institución organizadora:
Sociedad de Biofísica Argentina
Resumen:
Plasma membrane calcium pump (PMCA) is a P-ATPase that
transports Ca2+ against the electrochemical gradient from the cytoplasm
to the outer cellular medium. The principal modulator of PMCA is calmodulin which
increase its Ca2+ affinity and the maximum transport rate. The
reaction cycle of the pump can be described by the E1-E2 model.
The first step is the Ca2+ binding to the protein, drawing it to the
E1 conformation. The ATP binding and the later autophosphorylation elicit
PMCA to E1P. In the next step the pump reaches the E2P
conformation, which allows the liberation of Ca2+ to the cytoplasm.
However it has been described that besides the catalytic role of ATP; there is
another binding site for the nucleotide with a regulatory role associated with
the E2 conformation.
The aim of this work is to study the ATP binding to
purified PMCA and the associated conformational changes, using the fluorescent
analogue TNP-ATP. We also tested the conformational changes associated with the
presence of calmodulin.
Our results show that: (1) the TNP-ATP binds to the
PMCA (2) The environment of the ATP binding domain changes in the different
conformations of the PMCA. (3) The PMCA affinity for the TNP-ATP depends on the
PMCA conformation (4) The PMCA affinity for TNP-ATP is much lower in E2
conformation than in E1, suggesting the existence of an ATP low-affinity
binding site(s).
[1] Shmuel Muallem
and Steven J. D. Karlish.(1983) J.Biol.Chem. 258 159-175
[2] Suzuki, H., Kubota, T. ,Kubo ,K. ,Kanazawa, T.
(1990) Biochemistry. 29, 7040-7045
With grants of ANPCYT, CONICET, UBACYT y NIH.