-Ketoacids Are Potent Slow Binding Inhibitors of the Hepatitis C Virus NS3 Protease
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- 作者：Frank Narjes ; Mirko Brunetti ; Stefania Colarusso ; Benjamin Gerlach ; Uwe Koch ; Gabriella Biasiol ; Daniela Fattori ; Raffaele De Francesco ; Victor G. Matassa ; and Christian Steinkü ; hler
- 刊名：Biochemistry
- 出版年：2000
- 出版时间：February 22, 2000
- 年：2000
- 卷：39
- 期：7
- 页码：1849 - 1861
- 全文大小：272K
- 年卷期：v.39,no.7(February 22, 2000)
- ISSN：1520-4995
文摘
The replication of the hepatitis C virus (HCV), an important human pathogen, crucially dependson the proteolytic maturation of a large viral polyprotein precursor. The viral nonstructural protein 3(NS3) harbors a serine protease domain that plays a pivotal role in this process, being responsible forfour out of the five cleavage events that occur in the nonstructural region of the HCV polyprotein. Wehere show that hexapeptide, tetrapeptide, and tripeptide 
-ketoacids are potent, slow binding inhibitors ofthis enzyme. Their mechanism of inhibition involves the rapid formation of a noncovalent collision complexin a diffusion-limited, electrostatically driven association reaction followed by a slow isomerization stepresulting in a very tight complex. pH dependence experiments point to the protonated catalytic His 57 asan important determinant for formation of the collision complex. Ki values of the collision complexesvary between 3 nM and 18.5 
M and largely depend on contacts made by the peptide moiety of theinhibitors. Site-directed mutagenesis indicates that Lys 136 selectively participates in stabilization of thetight complex but not of the collision complex. A significant solvent isotope effect on the isomerizationrate constant is suggestive of a chemical step being rate limiting for tight complex formation. The potencyof these compounds is dominated by their slow dissociation rate constants, leading to complex half-livesof 11-48 h and overall Ki* values between 10 pM and 67 nM. The rate constants describing the formationand the dissociation of the tight complex are relatively independent of the peptide moiety and appear topredominantly reflect the intrinsic chemical reactivity of the ketoacid function.
-ketoacids are potent, slow binding inhibitors ofthis enzyme. Their mechanism of inhibition involves the rapid formation of a noncovalent collision complexin a diffusion-limited, electrostatically driven association reaction followed by a slow isomerization stepresulting in a very tight complex. pH dependence experiments point to the protonated catalytic His 57 asan important determinant for formation of the collision complex. Ki values of the collision complexesvary between 3 nM and 18.5 M and largely depend on contacts made by the peptide moiety of theinhibitors. Site-directed mutagenesis indicates that Lys 136 selectively participates in stabilization of thetight complex but not of the collision complex. A significant solvent isotope effect on the isomerizationrate constant is suggestive of a chemical step being rate limiting for tight complex formation. The potencyof these compounds is dominated by their slow dissociation rate constants, leading to complex half-livesof 11-48 h and overall Ki* values between 10 pM and 67 nM. The rate constants describing the formationand the dissociation of the tight complex are relatively independent of the peptide moiety and appear topredominantly reflect the intrinsic chemical reactivity of the ketoacid function.