Identification of a novel determinant for membrane association in hepatitis C virus nonstructural protein 4B
| Autor: | Valérie Castet, Hubert E. Blum, Jean Marie Ruysschaert, Darius Moradpour, François Penin, Vincent Raussens, Roland Montserret, Naveen Arora, Jérôme Gouttenoire, Eric Diesis |
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| Rok vydání: | 2009 |
| Předmět: |
Models
Molecular Magnetic Resonance Spectroscopy Viral protein Recombinant Fusion Proteins Green Fluorescent Proteins Molecular Sequence Data Immunology Hepacivirus Viral Nonstructural Proteins Biology medicine.disease_cause Microbiology Protein Structure Secondary Cell membrane 03 medical and health sciences Cell Line Tumor Virology Spectroscopy Fourier Transform Infrared medicine Humans Amino Acid Sequence Lipid bilayer Peptide sequence Integral membrane protein 030304 developmental biology 0303 health sciences 030306 microbiology Circular Dichroism Cell Membrane Transmembrane protein Virus-Cell Interactions Protein Structure Tertiary Transmembrane domain medicine.anatomical_structure Amino Acid Substitution Membrane protein Biochemistry Insect Science Sequence Alignment |
| Zdroj: | Journal of virology |
| DOI: | 10.1128/JVI.02663-08 |
| Popis: | Nonstructural protein 4B (NS4B) plays an essential role in the formation of the hepatitis C virus (HCV) replication complex. It is a relatively poorly characterized integral membrane protein predicted to comprise four transmembrane segments in its central portion. Here, we describe a novel determinant for membrane association represented by amino acids (aa) 40 to 69 in the N-terminal portion of NS4B. This segment was sufficient to target and tightly anchor the green fluorescent protein to cellular membranes, as assessed by fluorescence microscopy as well as membrane extraction and flotation analyses. Circular dichroism and nuclear magnetic resonance structural analyses showed that this segment comprises an amphipathic α-helix extending from aa 42 to 66. Attenuated total reflection infrared spectroscopy and glycosylation acceptor site tagging revealed that this amphipathic α-helix has the potential to traverse the phospholipid bilayer as a transmembrane segment, likely upon oligomerization. Alanine substitution of the fully conserved aromatic residues on the hydrophobic helix side abrogated membrane association of the segment comprising aa 40 to 69 and disrupted the formation of a functional replication complex. These results provide the first atomic resolution structure of an essential membrane-associated determinant of HCV NS4B. |
| Databáze: | OpenAIRE |
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