The Yeast Cell Fusion Protein Prm1p Requires Covalent Dimerization to Promote Membrane Fusion
The Yeast Cell Fusion Protein Prm1p Requires Covalent Dimerization to Promote Membrane Fusion
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DOI：
10.1371/journal.pone.0010593
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发表时间：
2010-05-11
期刊：
影响因子：
3.7
通讯作者：
Walter, Peter
中科院分区：
文献类型：
--
作者：
Engel, Alex;Aguilar, Pablo S.;Walter, Peter
Prm1p是一种多通道膜蛋白，在酵母交配过程中促进质膜融合。Prm1p和其他发育受控的细胞-细胞融合事件的假定调节因子促进膜融合的机制在很大程度上仍然难以捉摸。在这里，我们报告Prm1p形成共价连接的同源二聚体。共价Prm1p二聚体的形成是通过两个半胱氨酸Cys-120和Cys-545的分子间二硫键形成的。这些半胱氨酸被取代的PRM1突变体是融合缺陷的。这些PRM1突变体正常表达，保持同型相互作用，并可以运输到融合区。由于PRM1-C120S和PRM1-C545S突变体与野生型PRM1共表达时可以形成共价二聚体，因此推测分子间存在C120-C545二硫键。CyS-120与一个高度保守的疏水结构域相邻。这个疏水结构域中带电残基的突变会破坏共价二聚体的形成，转移到融合区，并促进融合的活性。分子间二硫键的重要性提供了关于PRM1介导的细胞-细胞融合机制的模型。
Prm1p is a multipass membrane protein that promotes plasma membrane fusion during yeast mating. The mechanism by which Prm1p and other putative regulators of developmentally controlled cell-cell fusion events facilitate membrane fusion has remained largely elusive. Here, we report that Prm1p forms covalently linked homodimers. Covalent Prm1p dimer formation occurs via intermolecular disulfide bonds of two cysteines, Cys-120 and Cys-545. PRM1 mutants in which these cysteines have been substituted are fusion defective. These PRM1 mutants are normally expressed, retain homotypic interaction and can traffic to the fusion zone. Because prm1-C120S and prm1-C545S mutants can form covalent dimers when coexpressed with wild-type PRM1, an intermolecular C120-C545 disulfide linkage is inferred. Cys-120 is adjacent to a highly conserved hydrophobic domain. Mutation of a charged residue within this hydrophobic domain abrogates formation of covalent dimers, trafficking to the fusion zone, and fusion-promoting activity. The importance of intermolecular disulfide bonding informs models regarding the mechanism of Prm1-mediated cell-cell fusion.