The study of the antigenic and sialic acid binding sites on the hemagglutinin molecule of influenza virus by recombinant DNA technique.

利用重组DNA技术研究流感病毒血凝素分子上的抗原和唾液酸结合位点。

• 批准号: 60480170
• 负责人: NAKAJIMA Katsuhisa
• 金额: $ 4.29万
• 依托单位: The Institute of Medical Science, The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1985
• 资助国家: 日本
• 起止时间: 1985 至 1986
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-60480170/
• 关键词: Influenza virus; Expression of HA protein; 部位突然変異法

Full-length cDNA of HA gene derived from B-1-23 virus, a variant of A/USSR/90/77 (A/USSR/77) (HlNl) which had been selected with monoclonal antibody (mAb) 264 and contained an amino acid change at position 190 of the HAl region, was cloned into the SV40 expression vector. Hemabsorbing or fusion activities of the HA protein expressed on CV-1 cells infected with the recombinant virus were indistinguishable from those of the authentic HA protein on B-1-23 virus infected cells. The antigenicity of the expressed HA protein was examined with five monoclonal antibodies to the HA of A/USSR/77 virus. The HA protein reacted with all mAbs except for mAb264. To define the area of epitope 264, we prepared seven HA cDNAs modified by site specific mutagenesis, and cloned them into the SV40 expression vector (SVHA). Raymond et al showed that changed amino acid residues 219 (lys) and 227 (Glu) in A/Brazil/11/78, and 189 (Lys) and 225 (Asp) in A/Lackland/3/78 were presumaly included in the area of epitope 265 by the nucleotide sequence analysis. Of those amino acids, our results shows it is the residues 190 and 219 and possibly residue 189 that are involved in the epitope 264, but neither the residue 225 nor 227.

将来自A/USSR/90/77（A/USSR/77）（H1N1）的变异株B-1-23病毒的HA基因的全长cDNA克隆到SV 40表达载体中。在用重组病毒感染的CV-1细胞上表达的HA蛋白的血吸收或融合活性与在B-1-23病毒感染的细胞上表达的真实HA蛋白的血吸收或融合活性没有区别。用5株抗A/USSR/77病毒HA的单克隆抗体检测表达的HA蛋白的抗原性。HA蛋白与除mAb 264之外的所有mAbs反应。为了确定表位264的区域，我们制备了七个通过位点特异性突变修饰的HA cDNA，并将它们克隆到SV 40表达载体（SVHA）中。Raymond等对A/Brazil/11/78和A/Lackland/3/78的219（Lys）和227（Glu）氨基酸残基和189（Lys）和225（Asp）氨基酸残基进行了核苷酸序列分析，推测它们位于表位265区域。在这些氨基酸中，我们的结果显示，残基190和219以及可能的残基189参与表位264，但残基225和227都不参与。

项目成果

Eri Nobusawa, Katsuhisa Nakajima and Setsuko Nakajima: "Expression of hemagglutinin gene" Vaccine. 3. 172-174 (1985)

Eri Nobusawa、Katsuhisa Nakajima 和 Settsuko Nakajima：“血凝素基因的表达”疫苗。

Eri Nobusawa;Katsuhisa Nakajima;Setsuko Nakajima: Virology.

Eri Nobusawa；Katsuhisa Nakajima；Settsuko Nakajima：病毒学。

Eri Nobusawa, Katsuhisa Nakajima and Setsuko Nakajima: "Determination of the epitope 264 on the hemagglutinin molecule of influenza HlNl virus by site specific mutagenesis." Virology. in press.

Eri Nobusawa、Katsuhisa Nakajima 和 Setsuko Nakajima：“通过位点特异性诱变测定流感 H1N1 病毒血凝素分子上的表位 264”。

Eri Nobusawa;Katsuhisa Nakajima;Setsuko Nakajima: Vaccine. 3. 172-174 (1985)

信泽绘里；中岛克久；中岛节子：疫苗。