title: The substrate recognition domain in the Na+/dicarboxylate and Na+/sulfate cotransporters is located in the carboxy-terminal portion of the protein.
abstract: The Na+/dicarboxylate cotransporter, NaDC-1, and the Na+/sulfate cotransporter, NaSi-1, share 43% sequence identity, but they exhibit no overlap in substrate specificity. A functional chimera, SiDC-4, was prepared from NaDC-1 and NaSi-1 by homologous recombination and expressed in Xenopus oocytes. SiDC-4 contains putative transmembrane domains 1-4 of NaSi-1 (amino acids 1-139) and putative transmembrane domains 5-11 of NaDC-1 (amino acids 141-593). SiDC-4 retains the substrate specificity of NaDC-1, which suggests that the substrate recognition domain is found in the carboxy-terminal portion of the protein, past amino acid 141. However, residues that affect substrate affinity and inhibition by furosemide and flufenamate are found in the amino terminal third of the protein. The cation binding properties of SiDC-4, including a stimulation of transport by lithium, differed from both parental transporters, suggesting that cation binding is determined by interactions between the amino- and carboxy-terminal portions of the protein. We conclude that the substrate recognition site of NaDC-1 and NaSi-1 is found in the carboxy-terminal portion of the protein, past amino acid 141, but residues in the amino terminus can affect substrate affinity, inhibitor sensitivity, and cation selectivity.
authors.affiliation: Department of Physiology, University of Arizona, College of Medicine, Tucson, AZ 85724, USA. Pajor@biosci.arizona.edu
authors.forename: A M
authors.initials: AM
authors.lastname: Pajor
authors.forename: N
authors.initials: N
authors.lastname: Sun
authors.forename: L
authors.initials: L
authors.lastname: Bai
authors.forename: D
authors.initials: D
authors.lastname: Markovich
authors.forename: P
authors.initials: P
authors.lastname: Sule
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000302:isolation & purification; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D002264:Carboxylic Acids / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D027682:Cation Transport Proteins; D002412:Cations; D029382:Dicarboxylic Acid Transporters; D017136:Ion Transport / Q000235:genetics; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics*; D009865:Oocytes; D029363:Organic Anion Transporters, Sodium-Dependent; D011995:Recombination, Genetic; D012964:Sodium / Q000378:metabolism*; D000077003:Sodium Sulfate Cotransporter; D013379:Substrate Specificity; D013431:Sulfates / Q000378:metabolism*; D027981:Symporters; D014981:Xenopus
chemical_list: D000906:Antibodies; D002264:Carboxylic Acids; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D002412:Cations; D029382:Dicarboxylic Acid Transporters; D008565:Membrane Proteins; D029363:Organic Anion Transporters, Sodium-Dependent; C092524:SLC13A2 protein, human; D000077003:Sodium Sulfate Cotransporter; D013431:Sulfates; D027981:Symporters; D012964:Sodium