EXPRESSION/DISTRIBUTION OF CONSERVED/SEROTYPE-SPECIFIC EPITOPES ON ROTAVIRUS VP8

轮状病毒 VP8 上保守/血清型特异性表位的表达/分布

• 批准号: 3803257
• 负责人: G LARRALDE
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3803257
• 关键词: Escherichia coli; Rotavirus; antibody neutralization test; antiserum; cross immunity; epitope mapping; genetic manipulation; guinea pigs; molecular cloning; plasmids; protein sequence; serotyping; virus antigen; virus classification; virus envelope; virus protein; western blottings

Three cDNA clones of approximately 300 nucleotides each and comprising the VP8 region of the VP4 protein of human rotavirus (HRV) strain KU, were constructed, inserted into the pGEMEX-1 plasmid and expressed in Escherichia coli. Immunoblot analysis of the corresponding peptides designated A (amino acid 1 to 105), B (amino acid 85 to 183), or C (amino acid 153 to 254), were performed using antisera to KU or Wa (VP4 serotype 1A), DS-1 (VP4 serotype 1B), or 1076 or M37 (VP4 serotype 2) rotavirus strains. These hyperimmune sera were prepared in guinea pigs inoculated with purified virus or expressed VP4 protein. The immunoblot analysis showed that epitopes located in fragment B were VP4 serotype and subtype specific and suggested that this fragment may be responsible for VP4 serotype and subtype specificity. In contrast, fragments A and C contained epitopes that were conserved among different human rotavirus VP4 serotypes. The high level of cross-reactivity observed in neutralization assay with antisera to the expressed VP8 protein may result from antibody to the conserved A and C fragments.

三个cDNA克隆，每个约300个核苷酸，包含 人轮状病毒（HRV）KU株的VP 4蛋白的VP 8区， 将其插入pGEMEX-1质粒，并在大肠杆菌中表达。 大肠埃希菌 相应肽的免疫印迹分析 命名为A（氨基酸1至105）、B（氨基酸85至183）或C （氨基酸153至254），使用KU或Wa（VP 4）的抗血清进行 血清型1A）、DS-1（VP 4血清型1B）或1076或M37（VP 4血清型2） 轮状病毒株。 这些超免疫血清在豚鼠中制备 接种纯化的病毒或表达的VP 4蛋白。免疫印迹 结果表明，片段B的抗原表位为VP 4血清型 和亚型特异性，并建议该片段可能负责 VP 4血清型和亚型特异性。 相反，片段A和 C含有在不同的人轮状病毒中保守的表位 VP 4血清型。 观察到的高水平交叉反应性， 用所表达的VP 8蛋白的抗血清进行的中和测定可 结果来自保守的A和C片段的抗体。