Increasing rate of cleavage at boundary between non-structural proteins 4B and 5A inhibits replication of Hepatitis C virus

Herod, M. R., Jones, D. M., McLauchlan, J. and McCormick, C. J. (2012) Increasing rate of cleavage at boundary between non-structural proteins 4B and 5A inhibits replication of Hepatitis C virus. Journal of Biological Chemistry, 287(1), pp. 568-580. (doi:10.1074/jbc.M111.311407) (PMID:22084249) (PMCID:PMC3249110)

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Abstract

In hepatitis C virus, non-structural proteins are cleaved from the viral polyprotein by viral encoded proteases. Although proteolytic processing goes to completion, the rate of cleavage differs between different boundaries, primarily due to the sequence at these positions. However, it is not known whether slow cleavage is important for viral replication or a consequence of restrictions on sequences that can be tolerated at the cleaved ends of non-structural proteins. To address this question, mutations were introduced into the NS4B side of the NS4B5A boundary, and their effect on replication and polyprotein processing was examined in the context of a subgenomic replicon. Single mutations that modestly increased the rate of boundary processing were phenotypically silent, but a double mutation, which further increased the rate of boundary cleavage, was lethal. Rescue experiments relying on viral RNA polymerase-induced error failed to identify second site compensatory mutations. Use of a replicon library with codon degeneracy did allow identification of second site compensatory mutations, some of which fell exclusively within the NS5A side of the boundary. These mutations slowed boundary cleavage and only enhanced replication in the context of the original lethal NS4B double mutation. Overall, the data indicate that slow cleavage of the NS4B5A boundary is important and identify a previously unrecognized role for NS4B5A-containing precursors requiring them to exist for a minimum finite period of time.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Jones, Dr Daniel and McLauchlan, Professor John
Authors: Herod, M. R., Jones, D. M., McLauchlan, J., and McCormick, C. J.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:Journal of Biological Chemistry
Journal Abbr.:J. Biol. Chem.
Publisher:American Society for Biochemistry and Molecular Biology, Inc.
ISSN:0021-9258
ISSN (Online):1083-351X
Published Online:14 November 2011

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
656341Virus-host interactions in hepatitis C virus infectionJohn MclauchlanMedical Research Council (MRC)MC_UU_12014/1MVLS III - CENTRE FOR VIRUS RESEARCH