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Journal of Virology, November 2009, p. 11078-11089, Vol. 83, No. 21
0022-538X/09/$08.00+0     doi:10.1128/JVI.00800-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Blocking Hepatitis C Virus Infection with Recombinant Form of Envelope Protein 2 Ectodomain{triangledown}

Jillian Whidby,1 Guaniri Mateu,2 Hannah Scarborough,2 Borries Demeler,3 Arash Grakoui,2 and Joseph Marcotrigiano1*

Center for Advanced Biotechnology and Medicine, Dept. of Chemistry and Chemical Biology, Rutgers University, 679 Hoes Lane, Piscataway, New Jersey 08854,1 Emory University School of Medicine, 954 Gatewood Road, NE, Atlanta, Georgia 30329,2 University of Texas, San Antonio, Dept. of Biochemistry, MC 7760, 7703 Floyd Curl Drive, San Antonio, Texas 78229-39013

Received 20 April 2009/ Accepted 13 August 2009

More than 120 million people worldwide are chronically infected with hepatitis C virus (HCV), making HCV infection the leading cause of liver transplantation in developed countries. Treatment is limited, and efficacy depends upon the infecting strain and the initial viral load. The HCV envelope glycoproteins (E1 and E2) are involved in receptor binding, virus-cell fusion, and entry into the host cell. HCV infection proceeds by endosomal acidification, suggesting that fusion of the viral envelope with cellular membranes is a pH-triggered event. E2 consists of an amino-terminal ectodomain, an amphipathic helix that forms a stem region, and a carboxy-terminal membrane-associating segment. We have devised a novel expression system for the production of a secreted form of E2 ectodomain (eE2) from mammalian cells and performed a comprehensive biochemical and biophysical characterization. eE2 is properly folded, as determined by binding to human CD81, blocking of infection of cell culture-derived HCV, and recognition by antibodies from patients chronically infected with different genotypes of HCV. The glycosylation pattern, number of disulfide bonds, oligomerization state, and secondary structure of eE2 have been characterized using mass spectrometry, size exclusion chromatography, circular dichroism, and analytical ultracentrifugation. These results advance the understanding of E2 and may assist in the design of an HCV vaccine and entry inhibitor.

* Corresponding author. Mailing address: Rutgers University, Center for Advanced Biotechnology and Medicine, Dept. of Chemistry and Chemical Biology, 679 Hoes Lane, Piscataway, NJ 08854. Phone: (732) 235-5309. Fax: (732) 235-5083. E-mail: jmarco{at}cabm.rutgers.edu

{triangledown} Published ahead of print on 26 August 2009.

Journal of Virology, November 2009, p. 11078-11089, Vol. 83, No. 21
0022-538X/09/$08.00+0     doi:10.1128/JVI.00800-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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