4AEP

HCV-JFH1 NS5B POLYMERASE STRUCTURE AT 1.8 ANGSTROM

Summary for 4AEP

• Entry DOI: 10.2210/pdb4aep/pdb
• Related: 2XXD 2XYM 4AEX
• Descriptor: RNA-DIRECTED RNA POLYMERASE, PHOSPHATE ION (3 entities in total)
• Functional Keywords: transferase, hepacivirus, nonstructural proteins, replication, de novo initiation, priming
• Biological source: HEPATITIS C VIRUS (HCV)
• Cellular location: Core protein p21: Host endoplasmic reticulum membrane; Single-pass membrane protein (By similarity). Core protein p19: Virion (By similarity). Envelope glycoprotein E1: Virion membrane; Single-pass type I membrane protein (Potential). Envelope glycoprotein E2: Virion membrane; Single-pass type I membrane protein (Potential). p7: Host endoplasmic reticulum membrane; Multi-pass membrane protein. Protease NS2-3: Host endoplasmic reticulum membrane; Multi-pass membrane protein (Potential). Serine protease NS3: Host endoplasmic reticulum membrane; Peripheral membrane protein (Probable). Non-structural protein 4A: Host endoplasmic reticulum membrane; Single-pass type I membrane protein (Potential). Non-structural protein 4B: Host endoplasmic reticulum membrane; Multi-pass membrane protein (By similarity). Non-structural protein 5A: Host endoplasmic reticulum membrane; Peripheral membrane protein (By similarity). RNA-directed RNA polymerase: Host endoplasmic reticulum membrane; Single-pass type I membrane protein (Potential): Q99IB8
• Total number of polymer chains: 1
• Total formula weight: 64821.09

Authors

Caillet-Saguy, C.,Bressanelli, S. (deposition date: 2012-01-12, release date: 2012-05-02, Last modification date: 2023-12-20)

Primary citation

Scrima, N.,Caillet-Saguy, C.,Ventura, M.,Harrus, D.,Astier-Gin, T.,Bressanelli, S.
Two Crucial Early Steps in RNA Synthesis by the Hepatitis C Virus Polymerase Involve a Dual Role of Residue 405.
J.Virol., 86:7107-, 2012

PubMed Abstract: The hepatitis C virus (HCV) NS5B protein is an RNA-dependent RNA polymerase essential for replication of the viral RNA genome. In vitro and presumably in vivo, NS5B initiates RNA synthesis by a de novo mechanism and then processively copies the whole RNA template. Dissections of de novo RNA synthesis by genotype 1 NS5B proteins previously established that there are two successive crucial steps in de novo initiation. The first is dinucleotide formation, which requires a closed conformation, and the second is the transition to elongation, which requires an opening of NS5B. We also recently published a combined structural and functional analysis of genotype 2 HCV-NS5B proteins (of strains JFH1 and J6) that established residue 405 as a key element in de novo RNA synthesis (P. Simister et al., J. Virol. 83:11926-11939, 2009; M. Schmitt et al., J. Virol 85:2565-2581, 2011). We hypothesized that this residue stabilizes a particularly closed conformation conducive to dinucleotide formation. Here we report similar in vitro dissections of de novo synthesis for J6 and JFH1 NS5B proteins, as well as for mutants at position 405 of several genotype 1 and 2 strains. Our results show that an isoleucine at position 405 can promote both dinucleotide formation and the transition to elongation. New structural results highlight a molecular switch of position 405 with long-range effects, resolving the implied paradox of how the same residue can successively favor both the closed conformation of the dinucleotide formation step and the opening necessary to the transition step.

PubMed: 22532694
DOI: 10.1128/JVI.00459-12

Experimental method

X-RAY DIFFRACTION (1.8 Å)