title: Helix packing of the cardiac Na+-Ca2+ exchanger: proximity of transmembrane segments 1, 2, and 6.
abstract: The cardiac Na+-Ca2+ exchanger (NCX1) is a membrane protein that extrudes Ca2+ from cells using the energy of the Na+ gradient and is a key protein in regulating intracellular Ca2+ and contractility. Based on the current topological model, NCX1 consists of nine transmembrane segments (TMSs). The N-terminal five TMSs are separated from the C-terminal four TMSs by a large intracellular loop. Cysteine 768 is modeled to be in TMS 6 close to the intracellular surface. In this study, the proximity of TMS 6 to TMSs 1 and 2 was examined. Insect High Five cells were transfected with cDNAs encoding mutant NCX1 proteins. Each mutant contained cysteine 768 and an introduced cysteine in TMS 1 or 2. Cross-linking between cysteines was determined after reaction with thiol-specific cross-linkers containing spacer arms of 6.5-12 A. The data indicate that residues in TMSs 1 and 2 are close to cysteine 768 in TMS 6. Cysteine 768 cross-linked with residues at both ends of TMSs 1 and 2 and is likely located toward the middle of TMS 6. Based on these results, we present an expanded helix-packing model for NCX1.
authors.affiliation: Department of Physiology, David Geffen School of Medicine at UCLA, Los Angeles, California 90095-1760, USA.
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Ren
authors.forename: Debora A
authors.initials: DA
authors.lastname: Nicoll
authors.forename: Kenneth D
authors.initials: KD
authors.lastname: Philipson
mesh_terms: D000818:Animals; D002118:Calcium / Q000737:chemistry; D002460:Cell Line; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003545:Cysteine / Q000737:chemistry; D004285:Dogs; D007313:Insecta; D009154:Mutation; D009206:Myocardium / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D019831:Sodium-Calcium Exchanger / Q000737:chemistry* / Q000235:genetics*
chemical_list: D003432:Cross-Linking Reagents; D019831:Sodium-Calcium Exchanger; C110374:sodium-calcium exchanger 1; D003545:Cysteine; D002118:Calcium