Metabolic Regulation of the Squid Nerve Na+/Ca2+ Exchanger: Recent Developments

  • Luis BeaugéEmail author
  • Reinaldo DiPolo
  • Mariana Bollo
  • Alexandra Cousido
  • Graciela Berberián
  • Alberto Podjarny
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 961)


In squid nerves, MgATP modulation of the Na+/Ca2+ exchanger requires the presence of a cytosolic protein which becomes phosphorylated during the process. This factor has been recently identified. Mass spectroscopy and Western blot analysis established that it is a member of the lipocalin superfamily of lipid-binding proteins (LBP or FABP) of 132 amino acids. We called it regulatory protein of squid nerve sodium/calcium exchanger (ReP1-NCXSQ, access to GenBank EU981897).

ReP1-NCXSQ was cloned, expressed, and purified. Circular dichroism, far-UV, and infrared spectroscopy suggest a secondary structure, predominantly of beta-sheets. The tertiary structure prediction provides ten beta-sheets and two alpha-helices, characteristic of most of LPB. Functional experiments showed that, to be active, ReP1-NCXSQ must be phosphorylated by MgATP, through the action of a kinase present in the plasma membrane. Moreover, PO4-ReP1-NCXSQ can stimulate the exchanger in the absence of ATP. An additional crucial observation was that, in proteoliposomes containing only the purified Na+/Ca2+ exchanger, PO4-ReP1-NCXSQ promotes activation; therefore, this upregulation has no other requirement than a lipid membrane and the incorporated exchanger protein.

Recently, we solved the crystal structure of ReP1-NCXSQ which was as predicted: a “barrel” consisting of ten beta-sheets and two alpha-helices. Inside the barrel is the fatty acid coordinated by hydrogen bonds with Arg126 and Tyr128. Point mutations showed that neither Tyr20Ala, Arg58Val, Ser99Ala, nor Arg126Val is necessary for protein phosphorylation or activity. On the other hand, Tyr128 is essential for activity but not for phosphorylation. We can conclude that (1) for the first time, a role of an LBP is demonstrated in the metabolic regulation of an ion exchanger; (2) phosphorylation of this LBP can be separated from the activation capacity; and (3) Tyr128, a candidate to coordinate lipid binding inside the barrel, is essential for activity.


Na+/Ca2+ exchanger Metabolic regulation Regulatory proteins Lipid-binding protein Structure-function Squid nerve Membrane transport 



The work was supported by grants from the US National Science Foundation (MCB 0444598); Fondo Nacional para Investigaciones Científicas y Tecnológicas (PICT-05-38073) and Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2010–2012: GI11220090100063), Argentina; Fondo Nacional para Ciencia y Técnica (S1-9900009046 and G-2001000637 FONACIT) and Fundación Polar, Venezuela; and CNRS and INSERM, Strasbourg, France.


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© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Luis Beaugé
    • 1
    Email author
  • Reinaldo DiPolo
    • 2
  • Mariana Bollo
    • 1
  • Alexandra Cousido
    • 3
  • Graciela Berberián
    • 1
  • Alberto Podjarny
    • 3
  1. 1.Laboratorio de BiofísicaInstituto de Investigación Médica “Mercedes y Martín Ferreyra” (INIMEC-CONICET)CórdobaArgentina
  2. 2.Laboratorio de Fisiología CelularCentro de Biofísica, Instituto Venezolano de Investigaciones Científicas (IVIC)CaracasVenezuela
  3. 3.Department of Structural Biology and Genomics, IGBMC, CNRS, INSERMUniversité de StrasbourgIllkirchFrance

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