Models to determine the kinetic mechanisms of ion- coupled transporters
Models to determine the kinetic mechanisms of ion- coupled transporters
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DOI：
10.1085/jgp.201812055
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发表时间：
2019-03-01
影响因子：
3.8
通讯作者：
Slotboom, Dirk J.
中科院分区：
文献类型：
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作者：
Lolkema, Juke S.;Slotboom, Dirk J.
随着高分辨率结构可用于许多离子耦合（次级活性）转运蛋白，该领域的一个主要挑战是确定如何实现耦合。运输反应的动力学机制的知识，它定义了基板和共离子的结合顺序，连同所有相关的状态被访问的转运蛋白的顺序，将有助于揭示这种耦合机制。在这里，我们推导出一般的数学模型，可用于分析稳态传输测量数据，并显示如何动力学机制可以得到。该模型描述了在不同的机制下，基质传输的表观最大速率如何取决于共离子浓度，反之亦然。类似地，它们描述了共离子浓度如何影响被运输底物的表观亲和力，反之亦然。最大速率和亲和力的分析允许扣除的前，一起，和后的基板结合的共离子的数量。希尔分析提供的信息较少，但在某些机制中，它可以揭示与基底一起运输的共离子的总数。然而，从其他实验方法的共离子的数量的先验知识是优选的，当推导动力学机制，因为模型通常是overparameterized。我们提出的模型具有广泛的适用性的离子耦合转运蛋白的研究。
With high-resolution structures available for many ion-coupled (secondary active) transporters, a major challenge for the field is to determine how coupling is accomplished. Knowledge of the kinetic mechanism of the transport reaction, which defines the binding order of substrate and co-ions, together with the sequence with which all relevant states are visited by the transporter, will help to reveal this coupling mechanism. Here, we derived general mathematical models that can be used to analyze data from steady-state transport measurements and show how kinetic mechanisms can be derived. The models describe how the apparent maximal rate of substrate transport depends on the co-ion concentration, and vice versa, in different mechanisms. Similarly, they describe how the apparent affinity for the transported substrate is affected by the co-ion concentration and vice versa. Analyses of maximal rates and affinities permit deduction of the number of co-ions that bind before, together with, and after the substrate. Hill analysis is less informative, but in some mechanisms, it can reveal the total number of co-ions transported with the substrate. However, prior knowledge of the number of co-ions from other experimental approaches is preferred when deriving kinetic mechanisms, because the models are generally overparameterized. The models we present have wide applicability for the study of ion-coupled transporters.