Orientation of Small Multidrug Resistance Transporter Subunits in the Membrane: Correlation with the Positive-Inside Rule
Orientation of Small Multidrug Resistance Transporter Subunits in the Membrane: Correlation with the Positive-Inside Rule
复制标题
DOI：
10.1016/j.jmb.2010.07.019
复制
发表时间：
2010-09-10
影响因子：
5.6
通讯作者：
Lolkema, Juke S.
中科院分区：
文献类型：
--
作者：
Kolbusz, Magdalena A.;ter Horst, Ramon;Lolkema, Juke S.
小型多药耐药（SMR）转运蛋白为较大的两结构域转运蛋白的进化提供了模型。使用报告基因融合技术确定了 SMR 家族 27 种蛋白质在膜中的方向。九个成员被单顺反子编码（单）并显示以两个方向插入（双拓扑）。十八个成员在染色体上成对编码，并显示以固定方向插入；每对中的两种蛋白质在膜中总是具有相反的方向。通过共纯化证明了两种蛋白质成对之间的相互作用。成对的蛋白质在膜中的方向仅受另一种蛋白质存在的轻微影响。对于成对的蛋白质，膜中的方向与细胞质和细胞外环上带正电荷的残基 (R + K) 的分布密切相关（正内规则）。相比之下，正内部规则对单链的双拓扑插入的预测效果较差。预计三个单链会以单一方向插入，N 末端和 C 末端位于膜的细胞外侧。电荷分布分析表明，细胞质环中的电荷数量需要达到阈值，内部正规则才具有预测价值。结论是，对染色体上的基因组织和系统发育进行组合分析足以区分 SMR 成员的固定或双重拓扑，以及可能相似类型的膜蛋白。正内规则可用于预测成对成员的方向，但不适合作为对偶拓扑的唯一预测器。 (C) 2010 Elsevier Ltd. 保留所有权利。
Small multidrug resistance (SMR) transport proteins provide a model for the evolution of larger two-domain transport proteins. The orientation in the membrane of 27 proteins from the SMR family was determined using the reporter fusion technique. Nine members were encoded monocistronically (singles) and shown to insert in both orientations (dual topology). Eighteen members were encoded in pairs on the chromosome and shown to insert in fixed orientations; the two proteins in each pair invariably had opposite orientations in the membrane. Interaction between the two proteins in pairs was demonstrated by copurification. The orientation in the membrane of either protein in the pair was affected only marginally by the presence of the other protein.For the proteins in pairs, the orientation in the membrane correlated well with the distribution of positively charges residues (R + K) over the cytoplasmic and extracellular loops (positive-inside rule). In contrast, dual-topology insertion of the singles was predicted less well by the positive-inside rule. Three singles were predicted to insert in a single orientation with the N-terminus and the C-terminus at the extracellular side of the membrane. Analysis of charge distributions suggests the requirement of a threshold number of charges in the cytoplasmic loops for the positive-inside rule to be of predictive value. It is concluded that a combined analysis of gene organization on the chromosome and phylogeny is sufficient to distinguish between fixed or dual topology of SMR members and, probably, similar types of membrane proteins. The positive-inside rule can be used to predict the orientation of members in pairs, but is not suitable as a sole predictor of dual topology. (C) 2010 Elsevier Ltd. All rights reserved.