The substrate specificity of a neuronal glutamate transporter is determined by the nature of the coupling ion.
Glutamate transporters are essential for terminating synaptic transmission. Glutamate is translocated together with three sodium ions. In the neuronal glutamate transporter EAAC1, lithium can replace sodium. To address the question of whether the coupling ion interacts with the 'driven' substrate during co-transport, the kinetic parameters of transport of the three substrates, L-glutamate and D- and L-aspartate by EAAC-1 in sodium- and lithium-containing media were compared. The major effect of the substitution of sodium by lithium was on Km. In the presence of sodium, the values for Km and Imax of these substrates were similar. In the presence of lithium, the Km for L-aspartate was increased around 13-fold. Remarkably, the corresponding increase for L-glutamate and D-aspartate was much larger, around 130-fold. In marked contrast, the Ki values for a non-transportable substrate analogue were similar in the presence of either sodium or lithium. The preference for L-aspartate in the presence of lithium was also observed when electrogenic transport of radioactive substrates was monitored in EAAC1-containing proteoliposomes. Our results indicate that, subsequent to substrate binding, the co-transported solutes interact functionally in the binding pocket of the transporter.
Menaker
David
D
Department of Biochemistry, Hebrew University Hadassah Medical School, Jerusalem, Israel.
Bendahan
Annie
A
Kanner
Baruch I
BI
D000600:Amino Acids, Dicarboxylic / Q000378:metabolism; D000818:Animals; D003470:Culture Media; D050580:Excitatory Amino Acid Transporter 3 / Q000235:genetics* / Q000378:metabolism*; D005260:Female; D007700:Kinetics; D023421:Models, Animal; D009474:Neurons / Q000378:metabolism*; D009865:Oocytes / Q000502:physiology; D018075:RNA, Complementary / Q000235:genetics; D011817:Rabbits; D013379:Substrate Specificity; D014158:Transcription, Genetic
D000600:Amino Acids, Dicarboxylic; D003470:Culture Media; D050580:Excitatory Amino Acid Transporter 3; D018075:RNA, Complementary