The hepatitis C virus NS3 proteinase: structure and function of a zinc-containing serine proteinase.
The hepatitis C virus (HCV) NS3 protein contains a serine proteinase domain implicated in the maturation of the viral polyprotein. NS3 forms a stable heterodimer with NS4A, a viral membrane protein that acts as an activator of the NS3 proteinase. The three-dimensional structure of the NS3 proteinase complexed with an NS4A-derived peptide has been determined. The NS3 proteinase adopts a chymotrypsin-like fold. A beta-strand contributed by NS4A is clamped between two beta-strands within the N terminus of NS3. Consistent with the requirement for extraordinarily long peptide substrates (P6-P4'), the structure of the NS3 proteinase reveals a very long, solvent-exposed substrate-binding site. The primary specificity pocket of the enzyme is shallow and closed at its bottom by Phe-154, explaining the preference of the NS3 proteinase for cysteine residues in the substrate P1 position. Another important feature of the NS3 proteinase is the presence of a tetrahedral zinc-binding site formed by residues Cys-97, Cys-99, Cys-145 and His-149. The zinc-binding site has a role in maintaining the structural stability and guiding the folding of the NS3 serine proteinase domain. Inhibition of the NS3 proteinase activity is regarded as a promising strategy to control the disease caused by HCV. Remarkably, the NS3 proteinase is susceptible to inhibition by the N-terminal cleavage products of substrate peptides corresponding to the NS4A/NS4B, NS4B/NS5A and NS5A/NS5B cleavage sites. The Ki values of the inhibitory products are lower than the K(m) values of the respective substrates and follow the order NS4A < NS5A < NS4B. Starting from the observation that the NS3 proteinase undergoes product inhibition, very potent, active site-directed inhibitors have been generated using a combinatorial peptide chemistry approach.
De Francesco
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Istituto di Ricerche di Biologia Molecolare P Angeletti, Pomezia, Rome, Italy. Defrancesco@IRBM.it
Pessi
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Steinkühler
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D000595:Amino Acid Sequence; D001665:Binding Sites / Q000187:drug effects; D004789:Enzyme Activation; D016174:Hepacivirus / Q000201:enzymology*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D020365:RNA Helicases; D012697:Serine Endopeptidases / Q000737:chemistry / Q000502:physiology*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013379:Substrate Specificity; D014759:Viral Envelope Proteins / Q000737:chemistry; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D015032:Zinc / Q000737:chemistry; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D020365:RNA Helicases; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors; D014759:Viral Envelope Proteins; D017361:Viral Nonstructural Proteins; D015032:Zinc; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C093542:NS4 protein, hepatitis C virus