A novel 13 kDa cytoplasmic soluble protein is required for the nucleotide (MgATP) modulation of the Na/Ca exchange in squid nerve fibers

• Published in: FEBS letters 1997-01, Vol.401 (1), p.6-10
• Main Authors: DiPolo, Reinaldo, Berberián, Graciela, Delgado, Daniel, Rojas, Hector, Beaugé, Luis
• Format: Article
• Language: English
• Subjects: 3-(N-morpholino) propanesulphonic acid; adenosine 5′-O-(thiotriphosphate); Adenosine Triphosphate - metabolism; Animals; ATPγS; Calcium - metabolism; calmodulin-dependent protein kinase; CAM-dependent PK; Carrier Proteins - metabolism; Cytoplasm - metabolism; Decapodiformes; EGTA; ethylene glycol-bis-β(aminoethyl ether)-N,N,N′,N′-tetraacetic acid; Loligo; Marine; Mops; N-methylglucamine; Na/Ca exchange regulation: MgATP; Nerve Fibers - metabolism; NMG; p-nitrophenylphosphate; p-NPP; PKA; PKC; protein kinase A; protein kinase C; Proteins - metabolism; PtyK; Sodium - metabolism; Sodium-Calcium Exchanger; Squid axon; tyrosine kinase
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(96)01416-0

The Na/Ca exchange is a highly regulated transport mechanism in which MgATP, a powerful modulatory intracellular substrate, has important implications for its function. As occurs with some preparations, in squid axons, nucleotide regulation is lost after membrane vesicle isolation. This has been a significant obstacle in the biochemical characterization of the MgATP effect. An important clue in solving this long-standing puzzle is presented in this work by showing that prolonged intracellular dialysis of squid axons produces a complete run down of the MgATP effect. Here we report that a soluble cytoplasmic factor isolated from fresh squid axoplasm and brain reconstitutes the MgATP stimulation of the Na-gradient-dependent 45Ca uptake in squid optic nerve membrane vesicles. Partial purification of this factor uncovers the presence of a novel 13 kDa soluble cytoplasmic protein (SCPr) which, when microinjected in ATP de-regulated dialyzed squid axons, completely restores the MgATP stimulation of Na o-dependent Ca efflux. We propose that in the squid preparation this SCPr constitutes the link between the nucleotide and target effector: the Na/Ca exchanger itself, or other plasma membrane structures which may secondarily interact with the exchanger.