A novel lipid binding protein is a factor required for MgATP stimulation of the squid nerve Na+/Ca2+ exchanger.
Here we identify a cytosolic factor essential for MgATP up-regulation of the squid nerve Na(+)/Ca(2+) exchanger. Mass spectroscopy and Western blot analysis established that this factor is a member of the lipocalin super family of lipid binding proteins of 132 amino acids in length. We named it Regulatory protein of the squid nerve sodium calcium exchanger (ReP1-NCXSQ). ReP-1-NCXSQ was cloned, over expressed and purified. Far-UV circular dichroism and infrared spectra suggest a majority of beta-strand in the secondary structure. Moreover, the predicted tertiary structure indicates ten beta-sheets and two short alpha-helices characteristic of most lipid binding proteins. Functional experiments showed that in order to be active ReP1-NCXSQ must become phosphorylated in the presence of MgATP by a kinase that is Staurosporin insensitive. Even more, the phosphorylated ReP1-NCXSQ is able to stimulate the exchanger in the absence of ATP. In addition to the identification of a new member of the lipid binding protein family, this work shows, for the first time, the requirement of a lipid binding protein for metabolic regulation of an ion transporting system.
Berberián
Graciela
G
Laboratorio de Biofísica, Instituto de Investigación Médica Mercedes y Martín Ferreyra (INIMEC-CONICET), Casilla de Correo 389, 5000 Córdoba, Argentina.
Bollo
Mariana
M
Montich
Guillermo
G
Roberts
Gretel
G
Degiorgis
Joseph A
JA
Dipolo
Reinaldo
R
Beaugé
Luis
L
D020121:5' Untranslated Regions / Q000235:genetics; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D049832:Decapodiformes / Q000502:physiology*; D005724:Ganglia / Q000502:physiology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009474:Neurons / Q000502:physiology*; D019831:Sodium-Calcium Exchanger / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D013055:Spectrophotometry, Infrared; D014466:Ultraviolet Rays
D020121:5' Untranslated Regions; D019831:Sodium-Calcium Exchanger; D000255:Adenosine Triphosphate