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Artículo

The role of the Ca2+ binding ligand Asn879 in the function of the plasma membrane Ca2+ pump

Rinaldi, Debora EugeniaIcon ; Adamo, Hugo PedroIcon
Fecha de publicación: 12/2009
Editorial: Elsevier Science
Revista: Biochimica et Biophysica Acta - Biomembranes
ISSN: 0005-2736
Idioma: Inglés
Tipo de recurso: Artículo publicado
Clasificación temática:
Bioquímica y Biología Molecular

Resumen

Asn879 in the transmembrane segment M6 of the plasma membrane Ca2+ pump (PMCA human isoform 4xb) has been proposed to coordinate Ca2+ at the transport site through its carboxylate. This idea agrees with the fact that this Asn is conserved in other Ca2+-ATPases but is replaced by Asp, Glu, and other residues in closely related 2P-type ATPases of different ionic specificity. Previous mutagenesis studies have shown that the substitution of Ala for Asn abolish the activity of the enzyme (Adebayo et al. 1995, Guerini et al. 1996). We have constructed a mutant PMCA in which the Asn879 was substituted by Asp. The mutant protein was expressed in S. cerevisiae,solubilized and purified by calmodulin affinity chromatography. The Asn879Asp PMCA mutant exhibited about 30 % of the wild type a Ca2+-dependent ATPase activity and only a minor reduction of the apparent affinity for Ca2+. The decrease in the Ca2+-ATPase of the mutant enzyme was in parallel with the reduction in the amount of phosphoenzyme formed from Ca2+ plus ATP. Noteworthy, the mutation nearly eliminated the ability of the enzyme to hydrolyze pNPP which is maximal in the absence of Ca2+ revealing a major effect of the mutation on the Ca2+-independent reactions of the transport cycle. At a pH low enough to protonate the Asp carboxylate the pNPPase activity of Asn879Asp increased, suggesting that the binding of protons to Asn879 is essential for the activities catalyzed by E2¬-like forms of the enzyme.
Palabras clave: ASPARAGINE , CA2+ BINDING SITE , CA2+ PUMP , MUTAGENESIS , P-ATPASE , PLASMA MEMBRANE CA2+-ATPASE , PMCA
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info:eu-repo/semantics/openAccess Excepto donde se diga explícitamente, este item se publica bajo la siguiente descripción: Creative Commons Attribution-NonCommercial-ShareAlike 2.5 Unported (CC BY-NC-SA 2.5)
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URI: http://hdl.handle.net/11336/138159
URL: https://www.sciencedirect.com/science/article/pii/S0005273609003058
DOI: http://dx.doi.org/10.1016/j.bbamem.2009.09.004
Colecciones
Articulos(IQUIFIB)
Articulos de INST.DE QUIMICA Y FISICO-QUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Citación
Rinaldi, Debora Eugenia; Adamo, Hugo Pedro; The role of the Ca2+ binding ligand Asn879 in the function of the plasma membrane Ca2+ pump; Elsevier Science; Biochimica et Biophysica Acta - Biomembranes; 1788; 11; 12-2009; 2404-2410
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