Expressed human hippocampal ASCT1 amino acid transporter exhibits a pH-dependent change in substrate specificity.
In mammalian cells, the basal Na+-dependent uptake for many of the neutral amino acids is mediated by a transport activity designated System ASC. A cloned human brain cDNA sequence, ASCT1, encodes a Na+-dependent neutral amino acid transport activity that exhibits a substrate specificity similar to that commonly associated with System ASC. However, the characteristics of ASC activity varies significantly between cell types and not all tissues contain detectable levels of ASCT1 mRNA. A unique property of System ASC activity is an altered substrate selectivity such that at pH values below 7.4 anionic amino acids function as inhibitors and substrates. The experiments in this report were designed to determine if the cloned ASCT1 transporter exhibited this pH-dependent anionic transport. Following transfection of HeLa cells with the ASCT1 cDNA, transport strongly favored neutral zwitterionic) amino acids when uptake was measured at a physiologic pH value of 7.5. However, lowering the assay pH to 5.5 significantly enhanced the interaction of the ASCT1 carrier with anionic amino acids such as cysteate, in a pH-dependent manner. The apparent pK for the titratable group was in the range of 6.5-7.0. These results provide evidence that the human brain ASCT1 transporter exhibits the most distinguishing characteristic known for System ASC and provides a model system to investigate the molecular basis for this shift in substrate acceptance.
Tamarappoo
B K
BK
Department of Biochemistry and Molecular Biology, University of Florida College of Medicine, Gainesville, 32610, USA.
McDonald
K K
KK
Kilberg
M S
MS
D026905:Amino Acid Transport Systems; D002352:Carrier Proteins / Q000032:analysis / Q000378:metabolism*; D002412:Cations / Q000378:metabolism; D003544:Cysteic Acid / Q000378:metabolism*; D006367:HeLa Cells; D006624:Hippocampus / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D012333:RNA, Messenger / Q000032:analysis; D012694:Serine / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D013379:Substrate Specificity; D014162:Transfection
D026905:Amino Acid Transport Systems; D002352:Carrier Proteins; D002412:Cations; D012333:RNA, Messenger; D012694:Serine; D012964:Sodium; D003544:Cysteic Acid