Lerch R A, Anderson K, Amann V L, Wertz G W
Department of Microbiology, University of Alabama Medical School, Birmingham 35294.
Virology. 1991 Mar;181(1):118-31. doi: 10.1016/0042-6822(91)90476-r.
Bovine respiratory syncytial (BRS) virus is an important cause of serious respiratory illness in calves. The disease caused in calves is similar to that caused by human respiratory syncytial (HRS) virus in children. The two viruses, however, have distinct host ranges and the attachment glycoproteins, G, have no antigenic cross-reactivity. The fusion glycoproteins, F, of the HRS and BRS viruses, however, have some antigenic cross-reactivity. To further compare the BRS virus and HRS virus fusion proteins, we determined the nucleotide sequence of cDNA clones to the BRS virus F protein mRNA, deduced the amino acid sequence, and compared these sequences with the HRS virus F protein sequences. The BRS virus F mRNA was 1899 nucleotides in length and had a single major open reading frame which could code for a polypeptide of 574 amino acids with an estimated molecular weight of 63.8 kDa. Structural features predicted from the amino acid sequence included an NH2-terminal signal sequence (residues 1-26), a site for proteolytic cleavage (residues 131-136) to generate the disulfide-linked F1 and F2 subunits, and a hydrophobic transmembrane anchor sequence (residues 522-549). The nucleic acid identity between the BRS virus and the HRS virus F mRNA sequences was 71.5%. The predicted BRS virus F protein shared 80.5% overall amino acid identity with the HRS virus F protein with 89% identity in the F1 polypeptide but only 68% identity in the F2 polypeptide. The position and number of the cysteine residues in the F1 and F2 polypeptides were conserved among all F proteins. However, BRS virus F protein had only three potential N-linked carbohydrate acceptor sites in comparison to four or five for the HRS viruses. A difference in the extent of glycosylation between the BRS and HRS virus F2 polypeptides was shown to be responsible for differences observed in the electrophoretic mobility of these proteins. A cDNA containing the complete open reading frame of the BRS virus F mRNA was inserted into the thymidine kinase gene of vaccinia virus and following homologous recombination, a recombinant virus containing the BRS virus F gene was isolated. The BRS virus F protein was expressed in recombinant virus infected cells as demonstrated by immunoprecipitation and was transported to and expressed on the surface of infected cells as shown by indirect immunofluorescence.
牛呼吸道合胞体（BRS）病毒是犊牛严重呼吸道疾病的重要病因。犊牛感染该病毒所引发的疾病与儿童感染人呼吸道合胞体（HRS）病毒所引发的疾病相似。然而，这两种病毒具有不同的宿主范围，其附着糖蛋白G不存在抗原交叉反应。不过，HRS病毒和BRS病毒的融合糖蛋白F具有一定的抗原交叉反应。为了进一步比较BRS病毒和HRS病毒的融合蛋白，我们测定了BRS病毒F蛋白mRNA的cDNA克隆的核苷酸序列，推导了氨基酸序列，并将这些序列与HRS病毒F蛋白序列进行了比较。BRS病毒F mRNA长度为1899个核苷酸，有一个单一的主要开放阅读框，可编码一个由574个氨基酸组成的多肽，估计分子量为63.8 kDa。从氨基酸序列预测的结构特征包括一个NH2末端信号序列（第1 - 26位残基）、一个用于蛋白水解切割以产生二硫键连接的F1和F2亚基的位点（第131 - 136位残基），以及一个疏水跨膜锚定序列（第522 - 549位残基）。BRS病毒和HRS病毒F mRNA序列之间的核酸同一性为71.5%。预测的BRS病毒F蛋白与HRS病毒F蛋白的总体氨基酸同一性为80.5%，在F1多肽中同一性为89%，但在F2多肽中仅为68%。F1和F2多肽中半胱氨酸残基的位置和数量在所有F蛋白中都是保守的。然而，与HRS病毒的四个或五个潜在N - 连接糖基化受体位点相比，BRS病毒F蛋白只有三个。BRS病毒和HRS病毒F2多肽糖基化程度的差异被证明是这些蛋白电泳迁移率观察到差异的原因。将包含BRS病毒F mRNA完整开放阅读框的cDNA插入痘苗病毒的胸苷激酶基因中，经过同源重组后，分离出含有BRS病毒F基因的重组病毒。如免疫沉淀所示，BRS病毒F蛋白在重组病毒感染的细胞中表达，如间接免疫荧光所示，它被转运到感染细胞表面并在其上表达。
