Vesicular Glutamate Transporters Use Flexible Anion and Cation Binding Sites for Efficient Accumulation of Neurotransmitter
Vesicular Glutamate Transporters Use Flexible Anion and Cation Binding Sites for Efficient Accumulation of Neurotransmitter
复制标题
DOI：
10.1016/j.neuron.2014.11.008
复制
发表时间：
2014-12-17
期刊：
影响因子：
16.2
通讯作者：
Jahn, Reinhard
中科院分区：
文献类型：
--
作者：
Preobraschenski, Julia;Zander, Johannes-Friedrich;Jahn, Reinhard
囊泡谷氨酸转运体（VGLUT）在突触囊泡中积累神经递质谷氨酸。运输依赖于V-ATP酶依赖的电化学质子梯度（Δ mu H+），并需要氯离子，但如何氯的行为和如何在运输过程中保持离子和电荷平衡是有争议的。使用重建方法，我们使用外源性质子泵来驱动VGLUT介导的转运，无论是在含有纯化VGLUT 1的脂质体中还是在与含有质子泵的脂质体融合的突触囊泡中。我们的数据表明，氯离子刺激可以在膜的两侧诱导。此外，氯化物在高浓度下与谷氨酸竞争。此外，VGLUT 1具有能够结合H+或K+离子的阳离子结合位点，允许质子反向转运或K+/H+交换。我们得出结论，VGLUT包含两个阴离子结合位点和一个阳离子结合位点，允许转运蛋白在囊泡填充过程中适应不断变化的离子条件，而不依赖于其他转运蛋白或通道。
Vesicular glutamate transporters (VGLUTs) accumulate the neurotransmitter glutamate in synaptic vesicles. Transport depends on a V-ATPase-dependent electrochemical proton gradient (Delta mu H+) and requires chloride ions, but how chloride acts and how ionic and charge balance is maintained during transport is controversial. Using a reconstitution approach, we used an exogenous proton pump to drive VGLUT-mediated transport either in liposomes containing purified VGLUT1 or in synaptic vesicles fused with proton-pump-containing liposomes. Our data show that chloride stimulation can be induced at both sides of the membrane. Moreover, chloride competes with glutamate at high concentrations. In addition, VGLUT1 possesses a cation binding site capable of binding H+ or K+ ions, allowing for proton antiport or K+/H+ exchange. We conclude that VGLUTs contain two anion binding sites and one cation binding site, allowing the transporter to adjust to the changing ionic conditions during vesicle filling without being dependent on other transporters or channels.