Cell Physiology and Molecular Biophysics, Texas Tech University Health Sciences Center, Lubbock, TX 79430.
Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, TX 79430.
Proc Natl Acad Sci U S A. 2019 Aug 20;116(34):16829-16834. doi: 10.1073/pnas.1901888116. Epub 2019 Aug 6.
Here, we present the atomic resolution crystallographic structure, the function, and the ion-binding properties of the KcsA mutants, G77A and G77C, that stabilize the 2,4-ion-bound configuration (i.e., water, K, water, K-ion-bound configuration) of the K channel's selectivity filter. A full functional and thermodynamic characterization of the G77A mutant revealed wild-type-like ion selectivity and apparent K-binding affinity, in addition to showing a lack of C-type inactivation gating and a marked reduction in its single-channel conductance. These structures validate, from a structural point of view, the notion that 2 isoenergetic ion-bound configurations coexist within a K channel's selectivity filter, which fully agrees with the water-K-ion-coupled transport detected by streaming potential measurements.
在这里，我们展示了 KcsA 突变体 G77A 和 G77C 的原子分辨率晶体结构、功能以及离子结合特性，这些突变体稳定了 K 通道选择性过滤器中的 2,4-离子结合构象（即水、K、水、K 离子结合构象）。对 G77A 突变体的全面功能和热力学特性进行了表征，结果表明其具有类似于野生型的离子选择性和明显的 K 结合亲和力，同时还表现出缺乏 C 型失活门控和其单通道电导明显降低的特点。这些结构从结构角度验证了这样一种观点，即在 K 通道的选择性过滤器中存在两种等能量的离子结合构象，这与流动电位测量所检测到的水-K 离子偶联运输完全一致。
