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. 2008 Sep 17;105(38):14545–14550. doi: 10.1073/pnas.0807298105

Fig. 5.

Fig. 5.

Mechanistic model of the membrane association process of NS3-4A. See Discussion for comments. The cytosolic side of the membrane bilayer is on the top. (1) N-terminal portion of NS3 (amino acids 13–23 highlighted in green) before folding of helix α0 (constructed by using the NS3 serine protease structure without the central segment of NS4A solved by X-ray crystallography (ref. 24; PDB entry 1A1Q). (2) Membrane association of uncleaved NS3-NS4A by NS3 helix α0. The NS4A structure is undefined at this step. It is represented as sticks with N-terminal segment 1–20, central segment 21–32, and the beginning of the C-terminal segment (amino acids 33–40) in medium, light, and dark orange, respectively. The NS3 structure was constructed by using: (i) The crystal structure of scNS3–4A (ref. 5; PDB entry 1CU1), where the C terminus of the helicase domain (silver) lies within the active site of the serine protease domain (cyan); (ii) The NS3 serine protease structure without the central segment of NS4A solved by NMR (ref. 25; PDB entry 1BT7) for the N-terminal β-barrel subdomain; and (iii) The NMR structure of NS3[10–24] comprising helix α0 (this study). Well folded structures (helix α0, C-terminal β-barrel subdomain, and helicase domain) are represented as ribbon diagrams whereas the less stable or unfolded structures are represented as sticks (NS3 segment 1–9 and N-terminal β-barrel subdomain). Side-chain atoms of the catalytic triad (His 57, Asp 81, and Ser 139) are highlighted as purple spheres. (3) Cleavage at the NS3/NS4A site allows membrane insertion of the N-terminal segment of NS4A, resulting in a transmembrane α-helix. (4) Cofolding of the central segment 21–32 of NS4A into the N-terminal β-barrel subdomain stabilizes the structure of the serine protease, which is locked onto the membrane by NS3 helix α0 and the NS4A transmembrane α-helix. Note that the hydrophilic helicase domain would be partially immersed into the membrane in the NS3-NS4A cis-cleavage conformation (5) (model in brackets). (5) Final topology of the NS3-4A complex on the membrane.