Hwang J, Fauzi H, Fukuda K, Sekiya S, Kakiuchi N, Shimotohno K, Taira K, Kusakabe I, Nishikawa S
National Institute of Bioscience & Human Technology, Tsukuba, Ibaraki, 305-8566, Japan.
Biochem Biophys Res Commun. 2000 Dec 20;279(2):557-62. doi: 10.1006/bbrc.2000.4007.
Nonstructural protein 3 (NS3) of hepatitis C virus (HCV) is a trypsin-like protease and is essential for processing of viral polyprotein. Accordingly, it is a potential target for anti-HCV drugs. Recently we could isolate RNA aptamers (G9-I, II, and III) which bind and inhibit NS3 protease using in vitro selection strategy. In addition, G9-I aptamer showed noncompetitive inhibition. In order to elucidate the binding site of G9-I aptamer in NS3 protease domain (deltaNS3), we carried out alanine scanning mutagenesis at positive charged residues on the surface of deltaNS3. The result of binding analysis by surface plasmon resonance measurements and protease inhibition assay clarified that Arg161 as well as Arg130 of deltaNS3 are essential for interaction with G9-I aptamer. This region appears to be a potential targeting site for anti-HCV drugs.
丙型肝炎病毒（HCV）的非结构蛋白3（NS3）是一种类胰蛋白酶，对病毒多聚蛋白的加工至关重要。因此，它是抗HCV药物的潜在靶点。最近，我们利用体外筛选策略分离出了能结合并抑制NS3蛋白酶的RNA适配体（G9-I、II和III）。此外，G9-I适配体表现出非竞争性抑制作用。为了阐明G9-I适配体在NS3蛋白酶结构域（deltaNS3）中的结合位点，我们对deltaNS3表面的带正电荷残基进行了丙氨酸扫描诱变。表面等离子体共振测量的结合分析结果和蛋白酶抑制试验表明，deltaNS3的Arg161以及Arg130对于与G9-I适配体的相互作用至关重要。该区域似乎是抗HCV药物的潜在靶向位点。
