Kim D W, Gwack Y, Han J H, Choe J
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Taejon, South Korea.
Virus Res. 1997 May;49(1):17-25. doi: 10.1016/s0168-1702(97)01452-4.
Hepatitis C virus (HCV) possesses two separate enzymatic functions in the NS3 protein: a protease and an NTPase/RNA helicase. In order to determine the minimal domain for NTPase and RNA helicase activities of the HCV NS3 protein, serial deletion mutants were constructed. The NS3H protein a fusion protein of 25 amino acids (aa) from an expression vector and the C-terminal 466 aa of the HCV NS3 protein, contains an NTPase/RNA helicase activity. We made deletion mutants of 10, 30, 50, 97, and 135 aa from the C-terminus and 16 and 32 aa from the N-terminus of the NS3H protein. The deleted protein lacking 50 aa from the C-terminus still possessed both activities, while the protein lacking 97 aa from the C-terminus lost an RNA helicase activity. The mutant lacking 16 amino acids from the N-terminus retained the enzymatic activities and the N-terminal 32 aa deleted mutant lost an NTPase/RNA helicase activity. A combinational deletion mutant lacking 16 aa the N-terminus and 50 aa from the C-terminus retained the enzymatic activities. These results show that the functional domain of the HCV NTPase/ RNA helicase is about 400 aa in length and maps between NS3 residues 1209 and 1608.
丙型肝炎病毒（HCV）的NS3蛋白具有两种独立的酶功能：一种蛋白酶和一种NTP酶/RNA解旋酶。为了确定HCV NS3蛋白NTP酶和RNA解旋酶活性的最小结构域，构建了一系列缺失突变体。NS3H蛋白是一种来自表达载体的25个氨基酸（aa）与HCV NS3蛋白C末端466个aa的融合蛋白，具有NTP酶/RNA解旋酶活性。我们从NS3H蛋白的C末端缺失了10、30、50、97和135个aa，从N末端缺失了16和32个aa。从C末端缺失50个aa的缺失蛋白仍具有这两种活性，而从C末端缺失97个aa的蛋白失去了RNA解旋酶活性。从N末端缺失16个氨基酸的突变体保留了酶活性，而N末端缺失32个aa的缺失突变体失去了NTP酶/RNA解旋酶活性。一个从N末端缺失16个aa且从C末端缺失50个aa的组合缺失突变体保留了酶活性。这些结果表明，HCV NTP酶/RNA解旋酶的功能结构域长度约为400个aa，位于NS3残基1209和1608之间。
