Site-directed mutagenesis reveals putative regions of protein interaction within the transmembrane domains of connexins.
Site-directed mutagenesis reveals putative regions of protein interaction within the transmembrane domains of connexins.
复制标题
定点诱变揭示了连接蛋白跨膜域内蛋白质相互作用的推定区域。
DOI：
10.1080/15419060802013463
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发表时间：
2008
影响因子：
--
通讯作者：
Skerrett,IM
中科院分区：
文献类型：
--
作者：
Toloue,MM;Woolwine,Y;Karcz,JA;Kasperek,EM;Nicholson,BJ;Skerrett,IM
通过半胱氨酸扫描诱变，作者比较了两个连接蛋白跨膜区域内的位点，一个来自α-类（Cx50），一个来自β-类（Cx32），其中氨基酸取代破坏了间隙连接通道的功能。在Cx32中，11个位点导致没有通道功能，或称为“反向门控”的异常电压门控表型，而在Cx50中，鉴定了7个这样的位点。在这两种连接蛋白中，位点都位于跨膜螺旋的特定面，这表明这些位点可能是跨膜结构域相互作用的位点。在Cx32中，M1跨膜结构域的一个宽面和M3的一个较窄的极性面被识别出来，包括一个被证明与封闭状态下的M4接近的位点。在Cx50中，发现了M3的相同面，但M1的敏感位点与Cx32不同。M1中较少的位点在Cx50中破坏通道功能，而那些被破坏的位点与Cx32中的敏感位点在不同的螺旋面上。对Cx32的M1和M2两个位点的更深入研究表明，侧链长度或分支对于维持正常的通道行为很重要，这与该位点是跨膜结构域相互作用的位点一致。
Through cysteine-scanning mutagenesis, the authors have compared sites within the transmembrane domains of two connexins, one from the α-class (Cx50) and one from the β-class (Cx32), where amino acid substitution disrupts the function of gap junction channels. In Cx32, 11 sites resulted in no channel function, or an aberrant voltage gating phenotype referred to as “reverse gating,” whereas in Cx50, 7 such sites were identified. In both connexins, the sites lie along specific faces of transmembrane helices, suggesting that these may be sites of transmembrane domain interactions. In Cx32, one broad face of the M1 transmembrane domain and a narrower, polar face of M3 were identified, including one site that was shown to come into close apposition with M4 in the closed state. In Cx50, the same face of M3 was identified, but sensitive sites in M1 differed from Cx32. Many fewer sites in M1 disrupted channel function in Cx50, and those that did were on a different helical face to the sensitive sites in Cx32. A more in depth study of two sites in M1 and M2 of Cx32 showed that side-chain length or branching are important for maintenance of normal channel behavior, consistent with this being a site of transmembrane domain interaction.