2XI3

HCV-H77 NS5B Polymerase Complexed With GTP

Summary for 2XI3

• Entry DOI: 10.2210/pdb2xi3/pdb
• Related: 1A1R 1A1V 1CWX 1HEI 1JR6 1N1L 1ONB 1R7C 1R7D 1R7E 1R7F 1R7G 1RGQ 2XI2
• Descriptor: RNA-directed RNA polymerase, GUANOSINE-5'-TRIPHOSPHATE, MAGNESIUM ION, ... (4 entities in total)
• Functional Keywords: transferase, nonstructural protein, replication, rdrp, de novo priming, hepacivirus
• Biological source: Hepatitis C virus (isolate H77)
• 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 (By similarity). 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. Non-structural protein 5A: Host endoplasmic reticulum membrane; Peripheral membrane protein. RNA-directed RNA polymerase: Host endoplasmic reticulum membrane; Single-pass type I membrane protein (Potential): P27958
• Total number of polymer chains: 2
• Total formula weight: 130309.73

Authors

Harrus, D.,Ahmed-El-Sayed, N.,Simister, P.C.,Miller, S.,Triconnet, M.,Hagedorn, C.H.,Mahias, K.,Rey, F.A.,Astier-Gin, T.,Bressanelli, S. (deposition date: 2010-06-25, release date: 2010-08-04, Last modification date: 2024-03-27)

Primary citation

Harrus, D.,Ahmed-El-Sayed, N.,Simister, P.C.,Miller, S.,Triconnet, M.,Hagedorn, C.H.,Mahias, K.,Rey, F.A.,Astier-Gin, T.,Bressanelli, S.
Further Insights Into the Roles of GTP and the C- Terminus of the Hepatitis C Virus Polymerase in the Initiation of RNA Synthesis
J.Biol.Chem., 285:32906-, 2010

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. Different structural elements of NS5b have been reported to participate in RNA synthesis, especially a so-called "β-flap" and a C-terminal segment (designated "linker") that connects the catalytic core of NS5b to a transmembrane anchor. High concentrations of GTP have also been shown to stimulate de novo RNA synthesis by HCV NS5b. Here we describe a combined structural and functional analysis of genotype 1 HCV-NS5b of strains H77 (subtype 1a), for which no structure has been previously reported, and J4 (subtype 1b). Our results highlight the linker as directly involved in lifting the first boundary to processive RNA synthesis, the formation of the first dinucleotide primer. The transition from this first dinucleotide primer state to processive RNA synthesis requires removal of the linker and of the β-flap with which it is shown to strongly interact in crystal structures of HCV NS5b. We find that GTP specifically stimulates this transition irrespective of its incorporation in neosynthesized RNA.

PubMed: 20729191
DOI: 10.1074/JBC.M110.151316

Experimental method

X-RAY DIFFRACTION (1.7 Å)