Visualizing ATP-dependent RNA translocation by the NS3 helicase from HCV

Area of Science:

• Virology
• Structural Biology
• Biochemistry

Background:

• Hepatitis C virus nonstructural protein 3 (NS3) is crucial for viral replication.
• NS3 is an ATP-dependent motor protein belonging to the superfamily 2 helicases.
• The precise mechanism of NS3 translocation along RNA is not fully understood.

Purpose of the Study:

• To determine the structural mechanism of hepatitis C virus NS3 translocation along RNA.
• To visualize the dynamic changes in NS3-RNA interaction during ATP hydrolysis.

Main Methods:

• Crystallographic analysis of full-length HCV NS3.
• Utilized bromine-labeled RNA oligonucleotides to track RNA positions.
• Determined structures of NS3 alone, NS3-RNA complex, and NS3-RNA-ATP analog complex.

Main Results:

• Observed large domain movements in NS3 upon ATP binding and hydrolysis.
• Demonstrated directional translocation of NS3 along RNA by one base pair per ATP hydrolyzed.
• Found the 3' end of RNA does not shift register, suggesting a 'spring-loading' mechanism.

Conclusions:

• ATP binding and hydrolysis induce domain movements enabling NS3 translocation along the RNA phosphodiester backbone.
• The 'spring-loading' mechanism explains how NS3 achieves larger translocation steps.
• Structural insights into NS3 function provide a basis for antiviral drug development.