Identification and characterization of a cDNA derived from multiple splicing that encodes envelope glycoprotein gp105 of human herpesvirus 6.
Identification and characterization of a cDNA derived from multiple splicing that encodes envelope glycoprotein gp105 of human herpesvirus 6.
复制标题
编码人疱疹病毒 6 型包膜糖蛋白 gp105 的多重剪接 cDNA 的鉴定和表征。
DOI：
10.1128/jvi.69.6.3490-3500.1995
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Chandran,B
中科院分区：
文献类型：
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作者：
Pfeiffer,B;Thomson,B;Chandran,B
糖蛋白复合物gp 82-gp 105是人疱疹病毒6型A型变异体（HHV-6A）的主要病毒体包膜糖蛋白复合物，由许多相关多肽组成。针对该糖蛋白复合物的单克隆抗体（MAb）2D 4、2D 6和13 D 6中和HHV-6A的感染性。我们先前已经报道了编码gp 82-gp 105复合物的病毒基因组DNA片段的一部分的分离、作图和表征以及中和表位的鉴定（B. Pfeiffer，Z. N. Berneman，F.内佩尔角K.张，S。Tirwatnapong和B. J. Virol. 67：4611-4620，1993）。该基因的进一步特征在于通过鉴定2.3-kb的基因组片段和通过鉴定2.5-kb的cDNA克隆。基因组序列含有编码由MAb识别的表位的短开放阅读框（ORF）。经Southern杂交分析，该cDNA对HHV-6具有特异性。在北方（RNA）印迹分析与总RNA从HHV-6A（GS）感染的细胞，cDNA插入特异性杂交与几个RNA种类。限制性酶切图谱分析将该cDNA定位于HHV-6A（U1102）基因组BamHI G片段，位于基因组独特长片段的右端，并分别定位于左末端和右末端同向重复区域内的SalI L和SalI O片段。在体外转录和翻译的cDNA揭示了约88.5 kDa的多肽，其在微粒体膜的存在下被糖基化为约104.2 kDa的多肽。两种多肽均通过MAb 2D 6进行免疫纯化，验证cDNA的身份为编码gp 82-gp 105复合物中的gp 105。cDNA序列分析显示，一个大的开放阅读框可能编码一个650个氨基酸的蛋白质与11个潜在的N-连接的糖基化位点和18个半胱氨酸残基。一个潜在的跨膜结构域仅位于推定蛋白的氨基末端附近，表明gp 105可能是2类糖蛋白。将cDNA核苷酸序列与来自HHV-6A（U1102）基因组BamH I G和SalI L片段的序列进行比较，发现编码gp 105的基因包含12个外显子，跨越病毒基因组的20 kb，内含子1跨越基因组DNA的约8 kb。cDNA的第一个外显子映射到左右末端的直接重复序列，而其他外显子映射到基因组的独特长片段内。（400字处截断摘要）
The glycoprotein complex gp82-gp105 is a major virion envelope glycoprotein complex of human herpesvirus 6 variant A (HHV-6A) and consists of a number of related polypeptides. Monoclonal antibodies (MAbs) 2D4, 2D6, and 13D6 against this glycoprotein complex neutralized HHV-6A infectivity. We have previously reported the isolation, mapping, and characterization of a portion of the viral genomic DNA fragment encoding the gp82-gp105 complex and the identification of the neutralizing epitope (B. Pfeiffer, Z. N. Berneman, F. Neipel, C. K. Chang, S. Tirwatnapong, and B. Chandran, J. Virol. 67:4611-4620, 1993). This gene was further characterized by the identification of a 2.3-kb genomic fragment and by the identification of a 2.5-kb cDNA clone. The genomic sequence contains a short open reading frame (ORF) encoding the epitope recognized by the MAbs. The identified cDNA showed specificity for HHV-6 in Southern blot analysis with viral DNA. In Northern (RNA) blot analysis with total RNA from HHV-6A(GS)-infected cells, the cDNA insert specifically hybridized with several RNA species. Restriction mapping analysis localized this cDNA to the HHV-6A(U1102) genomic BamHI G fragment, at the right end of the unique long segment of the genome and to the SalI L and SalI O fragments within the left and right terminal direct repeat regions, respectively. In vitro transcription and translation of the cDNA revealed a polypeptide of about 88.5 kDa which was glycosylated in the presence of microsomal membranes to a polypeptide of approximately 104.2 kDa. Both polypeptides were immunoprecipiated by MAb 2D6, verifying the identity of the cDNA as encoding the gp105 in the gp82-gp105 complex. Sequence analysis of the cDNA revealed a large ORF potentially encoding a 650-amino-acid protein with 11 potential N-linked glycosylation sites and 18 cysteine residues. A potential membrane-spanning domain is located only near the amino terminus of the putative protein, indicating that gp105 may be a class 2 glycoprotein. Comparison of the cDNA nucleotide sequence with sequences from HHV-6A(U1102) genomic BamHI G and SalI L fragments revealed that the gene encoding gp105 contains 12 exons, spanning over 20 kb of the viral genome, with intron 1 spanning about 8 kb of genomic DNA. The first exon of the cDNA mapped to the right and left terminal direct repeats, while the other exons mapped within the unique long segment of the genome.(ABSTRACT TRUNCATED AT 400 WORDS)