Viral replication. Structural basis for RNA replication by the hepatitis C virus polymerase.

Area of Science:

• Virology
• Structural Biology
• Drug Discovery

Background:

• Nucleotide analog inhibitors are clinically successful against Hepatitis C virus (HCV).
• The precise mechanism of the viral RNA-dependent RNA polymerase (NS5B) remains incompletely understood.
• Elucidating NS5B function is crucial for advancing HCV therapies.

Purpose of the Study:

• To determine high-resolution crystal structures of NS5B polymerase complexes.
• To elucidate the molecular mechanisms of HCV RNA replication initiation and elongation.
• To investigate interactions of nucleotide analog inhibitors within the NS5B active site.

Main Methods:

• X-ray crystallography of stalled NS5B polymerase ternary complexes.
• Complexes included RNA templates, primers, nucleotides, and metal ions.
• Analysis of structural changes during RNA synthesis.

Main Results:

• Identified conserved active-site residues in NS5B crucial for primer positioning.
• Described the dynamic role of a β loop and C-terminal linker in regulating active-site access.
• Revealed how these elements control initiation from the 3' terminus and subsequent elongation.
• Visualized the binding of nucleotide analog inhibitors, including sofosbuvir's active metabolite.

Conclusions:

• The study provides atomic-level insights into HCV RNA replication by NS5B.
• Structural dynamics of NS5B dictate replication initiation and elongation.
• Understanding these interactions facilitates the design of more effective HCV nucleotide analog inhibitors.