Early Intermediates in the Transport Cycle of the Neuronal Excitatory Amino Acid Carrier Eaac1
Early Intermediates in the Transport Cycle of the Neuronal Excitatory Amino Acid Carrier Eaac1
复制标题
神经元兴奋性氨基酸载体 Eaac1 运输周期的早期中间体
DOI：
--
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
C. Grewer
中科院分区：
文献类型：
--
作者：
N. Watzke;E. Bamberg;C. Grewer
兴奋性氨基酸载体1（EAAC 1）的产电谷氨酸转运与跨膜的多个电荷运动相关，这些电荷运动发生在微秒至毫秒的时间尺度上。这些电荷运动的分子性质目前知之甚少，因此，在本报告中详细研究了利用激光脉冲光解技术的笼谷氨酸提供了100 μs的时间分辨率。在内向传输模式中，谷氨酸诱导的耦合传输电流的瞬态分量的失活表现出两个指数分量。当通过去除钾将EAAC 1限制为Na+易位步骤时，获得了类似的结果，从而证明（1）EAAC 1的底物易位与正电荷的向内移动偶联，因此是产电的;以及（2）在EAAC 1转运循环的Na+结合和谷氨酸易位分支中存在至少两种不同的中间体。再加上确定的钠离子浓度和电压依赖性的两个指数的电荷运动和稳态EAAC 1的属性，我们开发了一个动力学模型，该模型是基于顺序结合的Na+和谷氨酸盐的细胞外结合位点EAAC 1解释我们的结果。在该模型中，至少一个Na+离子和随后的谷氨酸盐快速结合到转运蛋白，引发较慢的电中性结构变化，使EAAC 1能够进一步电压依赖性结合额外的钠离子。一旦形成完全负载的EAAC 1复合物，它可以经历慢得多的产电易位反应，以将底物和离子结合位点暴露于细胞质。
Electrogenic glutamate transport by the excitatory amino acid carrier 1 (EAAC1) is associated with multiple charge movements across the membrane that take place on time scales ranging from microseconds to milliseconds. The molecular nature of these charge movements is poorly understood at present and, therefore, was studied in this report in detail by using the technique of laser-pulse photolysis of caged glutamate providing a 100-μs time resolution. In the inward transport mode, the deactivation of the transient component of the glutamate-induced coupled transport current exhibits two exponential components. Similar results were obtained when restricting EAAC1 to Na+ translocation steps by removing potassium, thus, demonstrating (1) that substrate translocation of EAAC1 is coupled to inward movement of positive charge and, therefore, electrogenic; and (2) the existence of at least two distinct intermediates in the Na+-binding and glutamate translocation limb of the EAAC1 transport cycle. Together with the determination of the sodium ion concentration and voltage dependence of the two-exponential charge movement and of the steady-state EAAC1 properties, we developed a kinetic model that is based on sequential binding of Na+ and glutamate to their extracellular binding sites on EAAC1 explaining our results. In this model, at least one Na+ ion and thereafter glutamate rapidly bind to the transporter initiating a slower, electroneutral structural change that makes EAAC1 competent for further, voltage-dependent binding of additional sodium ion(s). Once the fully loaded EAAC1 complex is formed, it can undergo a much slower, electrogenic translocation reaction to expose the substrate and ion binding sites to the cytoplasm.
DOI：
10.1073/pnas.90.12.5767
发表时间：
1993-06-15
影响因子：
11.1
作者：
LOO, DDF;HAZAMA, A;WRIGHT, EM
通讯作者：
WRIGHT, EM
氢离子通过肾刷状缘谷氨酸转运蛋白协同转运。
影响因子：
2.9
作者：
Nelson,PJ;Dean,GE;Aronson,PS;Rudnick,G
通讯作者：
Rudnick,G