INVESTIGADORES
GONZALEZ FLECHA Francisco Luis
artículos
Título:
Phospholipid distribution around the plasma membrane calcium pump. A hydrophobic photolabeling study
Autor/es:
VILLAMIL GIRALDO A.M.; CASTELLO P.R.; GONZÁLEZ FLECHA F.L.; DELFINO J.M.; ROSSI J.P.F.C.
Revista:
Cell Biochemistry and Biophysics
Editorial:
Humana Press
Referencias:
Año: 2006 vol. 44 p. 431 - 437
ISSN:
1085-9195
Resumen:
The functions of membrane proteins are highly dependent on their
phospholipid environment. In this article, we have used a hydrophobic
photolabeling method to study the noncovalent interactions between
plasma membrane calcium pump (PMCA) and surrounding phospholipids. With
this approach, we determined (1) the number of lipid molecules in close
contact with the transmembrane surface, i.e., the lipid-protein
stoichiometry, and (2) the distribution of lipid molecules among
different regions of the protein. PMCA was photolabeled in mixed
micelles containing detergent, the phosphatidylcholine photoactivatable
analog 1-palmitoyl-2-[9-[2'-[125I]iodo-4'-
(trifluoromethyldiazirinyl)-benzyloxycarbonyl]-nonaoyl]-sn-glycero-3-phosphocholine,
and different amounts of dimyristoyl phosphatidylcholine (PC). The
stoichiometry was estimated after the extent of the labeling reaction
had been independently assessed. We determined a maximum number of 17
+/- 1 molecules of PC in close contact with the transmembrane surface
per PMCA molecule. In addition, a semiquantitative description of the
phospholipid environment around different regions of PMCA was carried
out after limited proteolysis of the photolabeled protein. The
distribution of labels among the N-terminal (1-322), the central
(323-660), and the C-terminal (661-1,205) regions was 26, 36, and 38%,
respectively.