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Journal of Virology, March 2007, p. 2745-2757, Vol. 81, No. 6
0022-538X/07/$08.00+0     doi:10.1128/JVI.01279-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Conserved Determinants for Membrane Association of Nonstructural Protein 5A from Hepatitis C Virus and Related Viruses{triangledown}

Volker Brass,1,{dagger} Zsuzsanna Pal,1,{dagger} Nicolas Sapay,2 Gilbert Deléage,2 Hubert E. Blum,1 François Penin,2 and Darius Moradpour1,3*

Department of Medicine II, University of Freiburg, D-79106 Freiburg, Germany,1 Institut de Biologie et Chimie des Protéines, UMR 5086, CNRS, Université de Lyon, IFR 128 BioSciences Lyon-Gerland, F-69397 Lyon, France,2 Division of Gastroenterology and Hepatology, Centre Hospitalier Universitaire Vaudois, University of Lausanne, CH-1011 Lausanne, Switzerland3

Received 18 June 2006/ Accepted 15 December 2006

Nonstructural protein 5A (NS5A) is a membrane-associated essential component of the hepatitis C virus (HCV) replication complex. An N-terminal amphipathic alpha helix mediates in-plane membrane association of HCV NS5A and at the same time is likely involved in specific protein-protein interactions required for the assembly of a functional replication complex. The aim of this study was to identify the determinants for membrane association of NS5A from the related GB viruses and pestiviruses. Although primary amino acid sequences differed considerably, putative membrane anchor domains with amphipathic features were predicted in the N-terminal domains of NS5A proteins from these viruses. Confocal laser scanning microscopy, as well as membrane flotation analyses, demonstrated that NS5As from GB virus B (GBV-B), GBV-C, and bovine viral diarrhea virus, the prototype pestivirus, display membrane association characteristics very similar to those of HCV NS5A. The N-terminal 27 to 33 amino acid residues of these NS5A proteins were sufficient for membrane association. Circular dichroism analyses confirmed the capacity of these segments to fold into alpha helices upon association with lipid-like molecules. Despite structural conservation, only very limited exchanges with sequences from related viruses were tolerated in the context of functional HCV RNA replication, suggesting virus-specific interactions of these segments. In conclusion, membrane association of NS5A by an N-terminal amphipathic alpha helix is a feature shared by HCV and related members of the family Flaviviridae. This observation points to conserved roles of the N-terminal amphipathic alpha helices of NS5A in replication complex formation.

* Corresponding author. Mailing address: Division of Gastroenterology and Hepatology, Centre Hospitalier Universitaire Vaudois, Rue du Bugnon 46, CH-1011 Lausanne, Switzerland. Phone: 41 21 314 47 23. Fax: 41 21 314 47 18. E-mail: Darius.Moradpour{at}chuv.ch.

{triangledown} Published ahead of print on 27 December 2006.

{dagger} V.B. and Z.P. contributed equally to this work.

Journal of Virology, March 2007, p. 2745-2757, Vol. 81, No. 6
0022-538X/07/$08.00+0     doi:10.1128/JVI.01279-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Mankouri, J., Milward, A., Pryde, K. R., Warter, L., Martin, A., Harris, M. (2008). A comparative cell biological analysis reveals only limited functional homology between the NS5A proteins of hepatitis C virus and GB virus B. J. Gen. Virol. 89: 1911-1920 [Abstract] [Full Text]  


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