Conformational cycle and ion-coupling mechanism of the Na+/hydantoin transporter Mhp1
Conformational cycle and ion-coupling mechanism of the Na+/hydantoin transporter Mhp1
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DOI：
10.1073/pnas.1410431111
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发表时间：
2014-10-14
影响因子：
11.1
通讯作者：
Mchaourab, Hassane S.
中科院分区：
文献类型：
--
作者：
Kazmier, Kelli;Sharma, Shruti;Mchaourab, Hassane S.
LeuT折叠的离子依赖性转运蛋白将生理必需分子的摄取与跨膜离子梯度偶联。由保守的5-螺旋反向重复序列定义，其编码离子和底物结合的共同原则，LeuT-折叠已被捕获在面向外、封闭和面向内的构象中。然而，有关交替访问和耦合到离子梯度的结构基础的基本问题仍然没有答案。在这里，我们使用的自旋标签对之间的距离测量来定义的Na+-耦合乙内酰脲同向转运体Mhp 1从微杆菌的构象循环。我们的研究结果表明，面向内和面向外的Mhp 1晶体结构代表采样的中间状态在溶液中。在这里，我们提供了一个机械的背景下，这些结构，将它们映射到一个模型的基础上的离子和底物依赖的构象平衡的运输。相反，Na+/亮氨酸转运蛋白LeuT，我们的研究结果表明，在保守的第二个Na+结合位点的Na+结合不改变能量的内向和外向的构象Mhp 1。LeuT和Mhp 1的配体依赖性交替访问的比较分析导致我们提出，不同的耦合方案的离子梯度可能会定义不同的构象机制内的LeuT折叠类。
Ion-dependent transporters of the LeuT-fold couple the uptake of physiologically essential molecules to transmembrane ion gradients. Defined by a conserved 5-helix inverted repeat that encodes common principles of ion and substrate binding, the LeuT-fold has been captured in outward-facing, occluded, and inward-facing conformations. However, fundamental questions relating to the structural basis of alternating access and coupling to ion gradients remain unanswered. Here, we used distance measurements between pairs of spin labels to define the conformational cycle of the Na+-coupled hydantoin symporter Mhp1 from Microbacterium liquefaciens. Our results reveal that the inward-facing and outward-facing Mhp1 crystal structures represent sampled intermediate states in solution. Here, we provide a mechanistic context for these structures, mapping them into a model of transport based on ion- and substrate-dependent conformational equilibria. In contrast to the Na+/leucine transporter LeuT, our results suggest that Na+ binding at the conserved second Na+ binding site does not change the energetics of the inward- and outward-facing conformations of Mhp1. Comparative analysis of ligand-dependent alternating access in LeuT and Mhp1 lead us to propose that different coupling schemes to ion gradients may define distinct conformational mechanisms within the LeuT-fold class.