uu.seUppsala University Publications
Change search
ReferencesLink to record
Permanent link

Direct link
Structural and Functional Analysis of Hepatitis C Virus Strain JFH1 Polymerase
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
Show others and affiliations
2009 (English)In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 83, no 22, 11926-11939 p.Article in journal (Refereed) Published
Abstract [en]

The hepatitis C virus (HCV) isolate JFH1 represents the only cloned wild-type sequence capable of efficient replication in cell culture, as well as in chimpanzees. Previous reports have pointed to the viral polymerase NS5B as a major determinant for efficient replication of this isolate. To understand the underlying mechanisms, we expressed and purified NS5B of JFH1 and of the closely related isolate J6, which replicates below the limit of detection in cell culture. The JFH1 enzyme exhibited a 5- to 10-fold-higher specific activity in vitro, consistent with the polymerase activity itself contributing to efficient replication of JFH1. The higher in vitro activity of the JFH1 enzyme was not due to increased RNA binding, elongation rate, or processivity of the polymerase but to higher initiation efficiency. By using homopolymeric and heteropolymeric templates, we found that purified JFH1 NS5B was significantly more efficient in de novo initiation of RNA synthesis than the J6 counterpart, particularly at low GTP concentrations, probably representing an important prerequisite for the rapid replication kinetics of JFH1. Furthermore, we solved the crystal structure of JFH1 NS5B, which displays a very closed conformation that is expected to facilitate de novo initiation. Structural analysis shows that this closed conformation is stabilized by a sprinkle of substitutions that together promote extra hydrophobic interactions between the subdomains "thumb" and "fingers." These analyses provide deeper insights into the initiation of HCV RNA synthesis and might help to establish more efficient cell culture models for HCV using alternative isolates.

Place, publisher, year, edition, pages
2009. Vol. 83, no 22, 11926-11939 p.
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-127451DOI: 10.1128/JVI.01008-09ISI: 000271084100050OAI: oai:DiVA.org:uu-127451DiVA: diva2:330077
Available from: 2010-07-14 Created: 2010-07-13 Last updated: 2010-08-26Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full text

Search in DiVA

By author/editor
Danielson, Helena
By organisation
Department of Biochemistry and Organic Chemistry
In the same journal
Journal of Virology
Medical and Health Sciences

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 417 hits
ReferencesLink to record
Permanent link

Direct link