Chao Y C, Chang M F, Gust I, Lai M M
Howard Hughes Medical Institute, University of Southern California, School of Medicine, Los Angeles 90033.
Virology. 1990 Oct;178(2):384-92. doi: 10.1016/0042-6822(90)90335-o.
The complete RNA sequence of the hepatitis delta virus (HDV) obtained from the Nauru Island in the Pacific was determined by cDNA cloning and amplification by polymerase chain reaction (PCR). The sequence showed 14-17% divergence from the two known HDV RNA sequences. There are three highly conserved domains: the region around the autocatalytic cleavage site of the genomic RNA (nucleotides 659 to 772), the region around the autocatalytic cleavage site of the antigenomic-sense RNA (nucleotides 847 to 966), and the region around the middle one-third domain of the open reading frame (ORF) encoding the hepatitis delta antigen on the antigenomic RNA (nucleotides 1267 to 1347). The two autocatalytic activities are required for the cleavage and ligation of HDV RNA during RNA replication. The third conserved domain codes for the RNA-binding domain of HDAg, which specifically interacts with HDV RNA. Three nucleotide changes within the genomic catalytic sequence are present but did not alter the catalytic cleavage activity of the HDV RNA. Microheterogeneity of the RNA sequences was also detected. One of these occurred within the coding region of the delta antigen, creating an amber termination codon in some of the RNA species. Thus, this HDV strain contains two different RNA species, one of which encodes a delta antigen of 214 amino acids and the other 195 amino acids. These two protein species were detected by immunoblotting of the patient's plasma. In contrast to other HDV strains, only three ORFs capable of encoding more than 100 amino acids each are present in this HDV RNA. We recommend that oligonucleotides complementary to the highly conserved sequences should be used as primers for PCR in clinical detection assays of hepatitis delta virus infection.
通过聚合酶链反应（PCR）进行cDNA克隆和扩增，测定了从太平洋瑙鲁岛获得的丁型肝炎病毒（HDV）的完整RNA序列。该序列与两个已知的HDV RNA序列有14%-17%的差异。有三个高度保守的结构域：基因组RNA自催化切割位点周围的区域（核苷酸659至772）、反基因组正义RNA自催化切割位点周围的区域（核苷酸847至966）以及反基因组RNA上编码丁型肝炎抗原的开放阅读框（ORF）中间三分之一结构域周围的区域（核苷酸1267至1347）。这两种自催化活性是HDV RNA在RNA复制过程中切割和连接所必需的。第三个保守结构域编码HDAg的RNA结合结构域，它与HDV RNA特异性相互作用。基因组催化序列内存在三个核苷酸变化，但未改变HDV RNA的催化切割活性。还检测到RNA序列的微异质性。其中之一发生在δ抗原的编码区内，在一些RNA种类中产生了一个琥珀终止密码子。因此，该HDV毒株包含两种不同的RNA种类，其中一种编码214个氨基酸的δ抗原，另一种编码195个氨基酸。通过对患者血浆进行免疫印迹检测到了这两种蛋白质种类。与其他HDV毒株不同，该HDV RNA中仅存在三个各自能够编码超过100个氨基酸的开放阅读框。我们建议，与高度保守序列互补的寡核苷酸应用作PCR引物，用于丁型肝炎病毒感染的临床检测试验。
