Alkhatib G, Roder J, Richardson C, Briedis D, Weinberg R, Smith D, Taylor J, Paoletti E, Shen S H
Division of Molecular Immunology and Neurobiology, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada.
J Virol. 1994 Oct;68(10):6770-4. doi: 10.1128/JVI.68.10.6770-6774.1994.
Membrane fusion caused by measles virus (MV) is a function of the fusion (F) protein. This process is essential for penetration into the host cell and subsequent initiation of the virus replicative cycle. The biological activity of the MV F protein is generated by endoproteolytic cleavage of a precursor protein (F0) into a large F1 subunit and a smaller F2 subunit held together by disulfide bonds. The cleavage site consists of a cluster of five basic amino acids (amino acids 108 to 112) within the predicted primary structure of the F protein. To investigate the role of the arginine residue at the carboxy terminus of the F2 subunit (arginine 112), site-directed mutagenesis was used to construct a cleavage mutant of the MV F protein in which this arginine residue was changed to a leucine residue. The mutated F gene, encoding four out of the five basic amino acids at the cleavage site, was inserted into the genome of vaccinia virus. The resulting recombinant virus was used to study expression of the mutant F protein in infected cells. Analysis of the Leu-112 mutant protein made in infected cells demonstrated that this single-amino-acid substitution resulted in a reduced rate of transport of the mutant protein to the cell surface, despite its efficient cleavage to yield F1 and F2 subunits. However, the electrophoretic mobilities of the Leu-112 polypeptides suggested that the protein was cleaved incorrectly. This aberrant cleavage appears to have abolished the ability of the F protein to cause syncytium formation. The data indicate that the arginine 112 residue is critical for the correct proteolytic cleavage that is required for the membrane fusion activity of the MV F protein.
麻疹病毒（MV）引起的膜融合是融合（F）蛋白的一项功能。这一过程对于病毒侵入宿主细胞并随后启动病毒复制周期至关重要。MV F蛋白的生物活性是通过将前体蛋白（F0）进行内蛋白水解切割，形成由二硫键连接在一起的一个较大的F1亚基和一个较小的F2亚基而产生的。切割位点由F蛋白预测一级结构内的一簇五个碱性氨基酸（氨基酸108至112）组成。为了研究F2亚基羧基末端的精氨酸残基（精氨酸112）的作用，采用定点诱变构建了MV F蛋白的一个切割突变体，其中该精氨酸残基被改变为亮氨酸残基。将编码切割位点五个碱性氨基酸中四个的突变F基因插入痘苗病毒基因组。所得重组病毒用于研究突变F蛋白在感染细胞中的表达。对感染细胞中产生的Leu-112突变蛋白的分析表明，尽管该突变蛋白能有效切割产生F1和F2亚基，但其向细胞表面的转运速率降低。然而，Leu-112多肽的电泳迁移率表明该蛋白切割错误。这种异常切割似乎消除了F蛋白引起合胞体形成的能力。数据表明，精氨酸112残基对于MV F蛋白膜融合活性所需的正确蛋白水解切割至关重要。
