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PDBsum entry 1r7c
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Membrane protein
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PDB id
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1r7c
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References listed in PDB file
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Key reference
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Title
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Structure and function of the membrane anchor domain of hepatitis c virus nonstructural protein 5a.
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Authors
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F.Penin,
V.Brass,
N.Appel,
S.Ramboarina,
R.Montserret,
D.Ficheux,
H.E.Blum,
R.Bartenschlager,
D.Moradpour.
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Ref.
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J Biol Chem, 2004,
279,
40835-40843.
[DOI no: ]
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PubMed id
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Abstract
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Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) is a
membrane-associated, essential component of the viral replication complex. Here,
we report the three-dimensional structure of the membrane anchor domain of NS5A
as determined by NMR spectroscopy. An alpha-helix extending from amino acid
residue 5 to 25 was observed in the presence of different membrane mimetic
media. This helix exhibited a hydrophobic, Trprich side embedded in detergent
micelles, while the polar, charged side was exposed to the solvent. Thus, the
NS5A membrane anchor domain forms an in-plane amphipathic alpha-helix embedded
in the cytosolic leaflet of the membrane bilayer. Interestingly, mutations
affecting the positioning of fully conserved residues located at the cytosolic
surface of the helix impaired HCV RNA replication without interfering with the
membrane association of NS5A. In conclusion, the NS5A membrane anchor domain
constitutes a unique platform that is likely involved in specific interactions
essential for the assembly of the HCV replication complex and that may represent
a novel target for antiviral intervention.
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Figure 5.
FIG. 5. NS5A membrane anchor mutants. A, amino acid
sequences of NS5A membrane anchor mutants. The amino acid
conservation among different HCV isolates (20) is indicated by
an asterisk (*), a colon (:), and a dot (.) for fully conserved,
conserved, and similar residues, respectively. B and C,
space-filling representation of theoretical helices for mutants
8+A and 11+A. An Ala insertion (shown in magenta) twists the
helix by 110°. The C-terminal side of the helix is shown in
the same orientation as in Fig. 3B to highlight the distortion
of charged residues on the N-terminal side. Residues are colored
as in Fig. 3B. These models were constructed by using the NMR
average structure of NS5A[1-31] observed in 100 mM SDS as
template and SwissPdb Viewer program (www.expasy.ch/swissmod/).
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Figure 8.
FIG. 8. Membrane association of NS5A mutants. Hypotonic
lysates of U-2 OS cells transiently transfected with
pCMVNS5Acon, -8+A, -11+A, and -  5-11 were analyzed by
centrifugation through Nycodenz gradients as described under
"Experimental Procedures." Fractions were collected from the top
and analyzed by immunoblot using mAbs 11H against NS5A and
G1/296 against p63.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2004,
279,
40835-40843)
copyright 2004.
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