Substrate specificity and functional characterization of sodium/dicarboxylate cotransporters

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Title: Substrate specificity and functional characterization of sodium/dicarboxylate cotransporters
Author: Naomi Oshiro
Abstract: Transport of dicarboxylates across the plasma membrane is mediated by the Na+ /dicarboxylate cotransporters (NaDCs ) belonging to the SLC13 gene family . These transporters play important roles in the homeostasis of dicarboxylates . The studies in this dissertation focused on two aspects of the NaDCs : structure -function studies of two low -affinity transporters , mouse (m ) and rabbit (rb ) NaDC1 , and functional characterization of a high -affinity NaDC transporter from Xenopus laevis , xNaDC3 . \r \n \r \nAlthough sharing strong sequence identity , mNaDC1 and rbNaDC1 differ in their ability to transport certain dicarboxylates . For example , oocytes expressing mNaDC1 exhibit large inward currents in the presence of glutarate , adipate , and succinate , whereas oocytes expressing rbNaDC1 have currents only with succinate . To identify NaDC1 domains involved in different ability to transport glutarate and adipate , I constructed a series of mNaDC1 -rbNaDC1 chimeras , and used both electrophysiological and dual -radiolabel competitive uptake techniques to exam their transport properties . My work indicates that different multiple transmembrane helices (TMs ) are involved in NaDC1 substrate recognition , with the region of TM 3 -4 and the C -terminus required for glutarate while the TM 8 -10 region is necessary for adipate transport . Further analysis of these two regions provided evidence that they contained residues important for both apparent substrate affinity and catalytic efficiency of NaDC1 . \r \n \r \nThe functional properties of non -mammalian vertebrates in the SLC13 family are not well characterized . Therefore , an initial functional characterization of xNaDC3 was performed using electrophysiological techniques . Like other members of the SLC13 family , xNaDC3 is electrogenic and exhibits inward substrate -dependent currents in the presence of sodium . However , other electrophysiological properties of xNaDC3 are unique and involve large cation -activated leak currents possibly mediated by anions . \r \n \r \nTaken together , these studies have provided insight into the mechanism of substrate recognition and transport by NaDCs . My work not only contributes to a more detailed analysis of NaDC structure -function relationships , but also demonstrates how transport protein structural information can be obtained using a biochemical approach . The need for such an approach can be explained by the fact that only a limited number of transporters have had their structures solved to an atomic resolution despite the critical involvement of transporters in cellular functions . \r \n
URI: http : / /hdl .handle .net /2152 .3 /201
Date: 2006-05-05


Substrate specificity and functional characterization of sodium/dicarboxylate cotransporters. Doctoral dissertation, The University of Texas Medical Branch. Available electronically from http : / /hdl .handle .net /2152 .3 /201 .

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