Structures of wild-type and a constitutively closed mutant of connexin26 shed light on channel regulation by CO 2
Structures of wild-type and a constitutively closed mutant of connexin26 shed light on channel regulation by CO 2
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connexin26 野生型和组成型封闭突变体的结构揭示了 CO 2 的通道调节
DOI：
10.1101/2023.08.22.554292
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Brotherton D
中科院分区：
文献类型：
--
作者：
Brotherton D
连接蛋白通过在并列的细胞之间形成缝隙连接通道(GJCs)来实现细胞间的通讯。连接蛋白26(Cx26)可直接受CO2调节，这可能是通过K125的氨甲酰化来实现的。我们发现，将K125突变为谷氨酸，模仿氨甲酰化的负电荷，导致Cx26 GJCs结构性关闭。通过低温EM，我们观察到K125E突变将构象平衡推向孔口狭窄的通道，类似于提高二氧化碳分压的效果。在以前的连接蛋白结构中，K125所在的胞质环处于无序状态，在调节中起重要作用。通过进一步的冷冻-EM研究，我们捕获了Cx26的不同状态，并观察了细胞质环的密度。这个环的位置、跨膜螺旋的构象和控制孔的N端螺旋的位置之间的相互作用，提供了一种调节机制。
Connexins allow intercellular communication by forming gap junction channels (GJCs) between juxtaposed cells. Connexin26 (Cx26) can be regulated directly by CO 2. This is proposed to be mediated through carbamylation of K125. We show that mutating K125 to glutamate, mimicking the negative charge of carbamylation, causes Cx26 GJCs to be constitutively closed. Through cryo-EM we observe that the K125E mutation pushes a conformational equilibrium towards the channel having a constricted pore entrance, similar to effects seen on raising the partial pressure of CO 2. In previous structures of connexins, the cytoplasmic loop, important in regulation and where K125 is located, is disordered. Through further cryo-EM studies we trap distinct states of Cx26 and observe density for the cytoplasmic loop. The interplay between the position of this loop, the conformations of the transmembrane helices and the position of the N-terminal helix, which controls the aperture to the pore, provides a mechanism for regulation.