Gating and Regulation of Connexin Hemichannels

20种人类连接蛋白同种型中的绝大多数与病理性 功能障碍，表明这些蛋白质在人类健康中的重要生理作用。 虽然连接蛋白通道对原子离子和小代谢物都是可渗透的，但它们是不可逆的。 对不同分子具有高度选择性，这可能决定了它们独特的生物学功能。 它们也受到各种刺激的调节，包括Ca 2+、电压和pH。 然而，渗透和门控的分子机制是非常肤浅的， 尤其是与其它经典离子通道相比时。最近，我们做出了令人惊讶的 发现连接蛋白半通道（和其他大孔通道）不是被动扩散 分子的孔隙。相反，它们显示饱和，表明渗透中的结合位点 可能以与转运蛋白/载体类似的方式决定选择性的途径。此外，本发明还 我们实验室发表的初步数据表明，N-末端（NT） 结构域，其内衬通道孔，在分子的渗透和孔闭塞。因此我们 假设连接蛋白半通道中的NT起以下作用：（1）作为一个关键的能量屏障， 分子的传输和（2）作为控制离子传导的门。在本提案中，我们 集中在两个连接蛋白，Cx 26和Cx 30，其中人类NT突变与 耳聋和先天性皮肤病。我们将确定这些NT突变是否影响 这可能会影响半通道的通透性、门控或两者，并将提供半通道功能障碍的机制基础。 连接蛋白通道的新的高分辨率结构已经解决，这将作为一个新的解决方案。 建议的分子动力学模拟指南。此外，我们将补充 计算数据与实验研究解决关键的机械问题， 还是没有答案我们的建议非常重要，因为我们正在测试一种创新的 这一假设将提供更好地理解渗透和门控的机制， 连接蛋白通道重要的是，这将有助于药物的合理开发和设计， 其他治疗方法，可以具体纠正或补偿不同的人类 连接蛋白功能障碍引起的疾病，包括耳聋和皮肤病。

The vast majority of the 20 human connexin isoforms have been associated with pathological dysfunctions, suggesting an essential physiological role for these proteins in human health. Although connexin channels are permeable to both atomic ions and small metabolites, they are highly selective to different molecules, which likely determines their distinct biological functions. They are also regulated by various stimuli, including Ca2+, voltage, and pH. Our understanding, however, of the molecular mechanisms underlying permeation and gating is extremely superficial, especially when compared with other classical ion channels. Recently, we made the surprising discovery that connexin hemichannels (and other large-pore channels) are not passive diffusion pores for molecules. Instead, they display saturation suggesting binding sites in the permeation pathway that might determine selectivity in a similar manner to transporters/carriers. Additionally, published and preliminary data from our lab suggested a major role for the N-terminal (NT) domain, which lines the channel pore, in permeation of molecules and pore occlusion. Thus, we hypothesize that the NT in connexin hemichannels serves: (1) as a critical energetic barrier for transport of molecules and (2) as the gate for controlling ionic conduction. In this proposal, we focus on two connexins, Cx26 and Cx30, in which human mutations at the NT are associated with deafness and congenital skin disorders. We will identify whether these NT mutations affect permeability, gating or both, and will provide a mechanistic basis of hemichannel dysfunction. Novel high-resolution structures of the connexin channel have been solved, which will serve as a guide for the proposed molecular dynamics simulations. Furthermore, we will complement computational data with experimental studies addressing key mechanistic questions that remain still unanswered. Our proposal is highly significant because we are testing an innovative hypothesis that will provide better understanding of the mechanisms of permeation and gating of connexin channels. Importantly, this will assist in the rational development and design of drugs or other therapeutic approaches that can specifically correct or compensate for the varied human diseases produced by connexin protein dysfunction, including deafness and skin disorders.