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J. Biol. Chem., Vol. 278, Issue 46, 45785-45792, November 14, 2003

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Unusual Multiple Recoding Events Leading to Alternative Forms of Hepatitis C Virus Core Protein from Genotype 1b^*

Steeve Boulant, Michel Becchi, François Penin, and Jean-Pierre Lavergne¶

From the Laboratoire de Bioinformatique et RMN Structurales and Centre Commun de Séquençage, Institut de Biologie et Chimie des Proteines, UMR5086 CNRS, Université Claude Bernard Lyon I, 7, Passage du Vercors, 69367 Lyon cedex 07, France

In addition to its involvement in the formation of the capsid shell of the virus particles, the core protein of hepatitis C virus (HCV) is believed to play an important role in the pathogenesis and/or establishment of persistent infection. We describe here alternative forms of genotype 1b HCV core protein identified after purification of various products of core protein segment 1-169 expressed in Escherichia coli and their analysis by proteolysis, mass spectrometry, and amino acid sequencing. These proteins all result from a +1 frameshift at codon 42 (a different position than that previously reported in genotype 1a) and, for some of them, from a rephasing in the normal open reading frame at the termination codon 144 in the +1 open reading frame. To test the relevance of these recoding events in a eukaryotic translational context, the nucleotide sequences surrounding the two shift sites were cloned in the three reading frames into expression vectors, allowing the production of a C-terminally fused green fluorescent protein, and expressed both in a reticulocyte lysate transcription/translation assay and in culture cells. Both recoding events were confirmed in these expression systems, strengthening the hypothesis that they might occur in HCV-infected cells. Moreover, sera from HCV-positive patients of genotype 1a or 1b were shown to react differently against synthetic peptides encoded in the +1 open reading frame. Together, these results indicate the occurrence of distinct recoding events in genotypes 1a and 1b, pointing out genotype-dependent specific features for F protein.

Received for publication, July 4, 2003 , and in revised form, September 2, 2003.

An oral presentation of part of this work was done at the 9^th International Meeting on Hepatitis C and Related Viruses (San Diego, CA, July, 2002).

^* This work was supported by a research grants from Université Claude Bernard Lyon I and from CNRS "Programme Physique et Chimie du Vivant,"Association pour la Recherche sur le Cancer Grant 4301 (to J.-P. L.), and "Programme de Recherche Fondamentale en Microbiologie et Maladies Infectieuses et Parasitaires" Ministere de l'Education Nationale, de la Recherche et des Technologies Grant 1A031G (to J.-P. L.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ To whom correspondence should be addressed. Tel.: 33-472-722-645; Fax: 33-472-722-605; E-mail: jp.lavergne{at}ibcp.fr.

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