Specific targeting of hepatitis C virus core protein by an intracellular single-chain antibody of human origin.
Specific targeting of hepatitis C virus core protein by an intracellular single-chain antibody of human origin.
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DOI：
10.1002/hep.22366
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发表时间：
2008-09
期刊：
影响因子：
13.5
通讯作者：
Heintges, Tobias
中科院分区：
文献类型：
--
作者：
Karthe, Juliane;Tessmann, Kathi;Li, Jisu;Machida, Raiki;Daleman, Maaike;Haeussinger, Dieter;Heintges, Tobias
丙型肝炎病毒 (HCV) 核心蛋白对于病毒基因组衣壳化至关重要，并在脂肪变性、免疫逃避和肝细胞癌中发挥重要作用。因此，它可能是干扰 HCV 生命周期和相关发病机制的一个有前途的治疗靶点。在本研究中，我们利用噬菌体展示技术从慢性丙型肝炎患者的骨髓浆细胞中产生针对核心蛋白的单链可变域抗体片段（scFv）。从而鉴定出具有高亲和力结合的抗体（scFv42C），并将结合位点定位到不同基因型之间保守的核心蛋白的PLXG基序（残基84-87）。 scFv42C 在 Huh7 人肝癌细胞系中单独表达时显示出弥散的细胞质荧光，而与核心基因共转染则将其亚细胞分布转变为核心蛋白的亚细胞分布。通过荧光共振能量转移技术独立证明了 scFv42C 与其靶核心蛋白的细胞内关联。有趣的是，单链抗体的表达降低了细胞内核心蛋白水平，特别是在丙型肝炎病毒完全复制的情况下。此外，核心蛋白诱导的细胞增殖可以通过 scFv4C 共表达来逆转。因此，scFv42C 可能代表一种新型抗 HCV 药物，其通过隔离核心蛋白并减弱核心蛋白介导的发病机制发挥作用。
The hepatitis C virus (HCV) core protein is essential for viral genome encapsidation and plays an important role in steatosis, immune evasion, and hepatocellular carcinoma. It may thus represent a promising therapeutic target to interfere with the HCV life-cycle and related pathogenesis. In this study, we used phage display to generate single-chain variable domain antibody fragments (scFv) to the core protein from bone marrow plasma cells of patients with chronic hepatitis C. An antibody with high-affinity binding (scFv42C) was thus identified, and the binding site was mapped to the PLXG motif (residues 84–87) of the core protein conserved among different genotypes. Whereas scFv42C displayed diffuse cytoplasmic fluorescence when expressed alone in the Huh7 human hepatoma cell line, cotransfection with the core gene shifted its subcellular distribution into that of core protein. The intracellular association of scFv42C with its target core protein was independently demonstrated by the fluorescence resonance energy transfer technique. Interestingly, expression of the single-chain antibody reduced core protein levels intracellularly, particularly in the context of full HCV replication. Moreover, cell proliferation as induced by the core protein could be reversed by scFv4C coexpression. Therefore, scFv42C may represent a novel anti-HCV agent, which acts by sequestering core protein and attenuating core protein–mediated pathogenesis.
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影响因子：
13.5
作者：
Fukutomi, T;Zhou, YH;Li, JS
通讯作者：
Li, JS
影响因子：
4.8
作者：
Goncalves, J;Silva, F;Barbas, C
通讯作者：
Barbas, C
影响因子：
4.8
作者：
Moradpour, D;Bieck, E;Bartenschlager, R
通讯作者：
Bartenschlager, R
影响因子：
2.2
作者：
Marasco, WA;LaVecchio, J;Winkler, A
通讯作者：
Winkler, A
DOI：
10.1073/pnas.91.13.5932
发表时间：
1994-06-21
影响因子：
11.1
作者：
CHEN, SY;KHOURI, Y;MARASCO, WA
通讯作者：
MARASCO, WA