Three conformational snapshots of the hepatitis C virus NS3 helicase reveal a ratchet translocation mechanism

被引:186
作者
Gu, Meigang [1 ]
Rice, Charles M. [1 ]
机构
[1] Rockefeller Univ, Ctr Study Hepatitis C, Lab Virol & Infect Dis, New York, NY 10065 USA
关键词
antivirals; motor protein; step size; superfamily; 2; transition state; RNA HELICASE; MUTATIONAL ANALYSIS; NUCLEOSIDE TRIPHOSPHATASE; CRYSTAL-STRUCTURE; STRUCTURAL BASIS; PROTEASE DOMAIN; DNA; COMPLEX; ATP; REPLICATION;
D O I
10.1073/pnas.0913380107
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
A virally encoded superfamily-2 (SF2) helicase (NS3h) is essential for the replication of hepatitis C virus, a leading cause of liver disease worldwide. Efforts to elucidate the function of NS3h and to develop inhibitors against it, however, have been hampered by limited understanding of its molecular mechanism. Here we show x-ray crystal structures for a set of NS3h complexes, including ground-state and transition-state ternary complexes captured with ATP mimics (ADP center dot BeF(3) and ADP center dot AlF(4)(-)). These structures provide, for the first time, three conformational snapshots demonstrating the molecular basis of action for a SF2 helicase. Upon nucleotide binding, overall domain rotation along with structural transitions in motif V and the bound DNA leads to the release of one base from the substrate base-stacking row and the loss of several interactions between NS3h and the 3' DNA segment. As nucleotide hydrolysis proceeds into the transition state, stretching of a "spring" helix and another overall conformational change couples rearrangement of the (d) NTPase active site to additional hydrogen-bonding between NS3h and DNA. Together with biochemistry, these results demonstrate a "ratchet" mechanism involved in the unidirectional translocation and define the step size of NS3h as one base per nucleotide hydrolysis cycle. These findings suggest feasible strategies for developing specific inhibitors to block the action of this attractive, yet largely unexplored drug target.
引用
收藏
页码:521 / 528
页数:8
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