Aminoterminal Amphipathic α-Helix AH1 of Hepatitis C Virus Nonstructural Protein 4B Possesses a Dual Role in RNA Replication and Virus Production
Figure 2
(A) Far UV circular dichroism (CD) spectra of synthetic peptide NS4B[1–40] recorded in 5 mM sodium phosphate pH 7.5 (H2O), complemented with either 50% 2,2,2-trifluoroethanol (TFE) or 1% L-α-lysophosphatidylcholine (LPC) or the following detergents: 100 mM sodium dodecyl sulfate (SDS), 100 mM n-dodecyl-β-D-maltoside (DDM), or 100 mM dodecylphosphocholine (DPC). (B) Summary of sequential (i, i+1) and medium-range (i, i+2 to i, i+4) nuclear Overhauser enhancements (NOEs) of NS4B[1–40] in 50% TFE. Sequential NOEs allowing the assignment of proline residues are indicated in red. Asterisks indicate that the presence of a NOE cross peak was not confirmed because of overlapping resonances or the lack of H assignment. Intensities of NOEs are indicated by the height of the bars. Amide protons that remained observable in nuclear magnetic resonance (NMR) spectra after three days in 50% D20/50% deuterated TFE (TFE-d3) are indicated by blue squares (slow exchangeable protons). (C) NMR-derived 1Hα and 13Cα chemical shift differences were calculated by subtraction of the experimental values from the reported random coil conformation values in TFE [61]. The dashed lines indicate the standard threshold value of ΔHα (−0.1 ppm) or ΔCα (0.7 ppm) for an α-helix. (D) Representative NMR structure of NS4B[1–40] peptide in 50% TFE. The aa sequence of NS4B[1–40] is depicted in (a). The box indicates α-helix residues. Residues are color-coded as described in the Legend to Figure 1. (b–d) representative NMR structure model of NS4B[1–40] (PDB entry 2LVG) showing the amphipathic character of α-helix 4–32. b, ribbon representation of the side view colored as in panel a; c and d, hydrophilic side and hydrophobic side views of the molecular surface of amphipathic α-helix 4–32. Figures were generated from structure coordinates using VMD (http://www.ks.uiuc.edu/Research/vmd/) [68] and rendered with POV-Ray (http://www.povray.org/).