Published:
American Society for Microbiology, 2007
Published in:
Journal of Virology, 81 (2007) 6, Seite 2745-2757
Language:
English
DOI:
10.1128/jvi.01279-06
ISSN:
0022-538X;
1098-5514
Origination:
Footnote:
Description:
ABSTRACT Nonstructural protein 5A (NS5A) is a membrane-associated essential component of the hepatitis C virus (HCV) replication complex. An N-terminal amphipathicalpha helix mediates in-plane membrane association of HCV NS5A and atthe same time is likely involved in specific protein-proteininteractions required for the assembly of a functional replicationcomplex. The aim of this study was to identify the determinants formembrane association of NS5A from the related GB viruses andpestiviruses. Although primary amino acid sequences differedconsiderably, putative membrane anchor domains with amphipathicfeatures were predicted in the N-terminal domains of NS5A proteins fromthese viruses. Confocal laser scanning microscopy, as well as membraneflotation 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 ofHCV NS5A. The N-terminal 27 to 33 amino acid residues of these NS5Aproteins were sufficient for membrane association. Circular dichroismanalyses confirmed the capacity of these segments to fold into alphahelices upon association with lipid-like molecules. Despite structuralconservation, only very limited exchanges with sequences from relatedviruses were tolerated in the context of functional HCV RNAreplication, suggesting virus-specific interactions of these segments.In conclusion, membrane association of NS5A by an N-terminalamphipathic alpha helix is a feature shared by HCV and related membersof the family Flaviviridae . This observation points toconserved roles of the N-terminal amphipathic alpha helices of NS5A inreplication complexformation.