Potent Peptide Inhibitors of Human Hepatitis C Virus NS3 Protease Are Obtained by Optimizing the Cleavage Products
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- 作者：Paolo Ingallinella ; Sergio Altamura ; Elisabetta Bianchi ; Marina Taliani ; Raffaele Ingenito ; Riccardo Cortese ; Raffaele De Francesco ; Christian Steinkü ; hler ; and Antonello Pessi
- 刊名：Biochemistry
- 出版年：1998
- 出版时间：June 23, 1998
- 年：1998
- 卷：37
- 期：25
- 页码：8906 - 8914
- 全文大小：259K
- 年卷期：v.37,no.25(June 23, 1998)
- ISSN：1520-4995
文摘
In the absence of a broadly effective cure for hepatitis caused byhepatitis C virus (HCV),much effort is currently devoted to the search for inhibitors of thevirally encoded protease NS3. Thischymotrypsin-like serine protease is required for the maturation of theviral polyprotein, cleaving it at theNS3-NS4A, NS4A-NS4B, NS4B-NS5A, and NS5A-NS5B sites. Inthe course of our studies on thesubstrate specificity of NS3, we found that the products of cleavagecorresponding to the P6-P1 regionof the substrates act as competitive inhibitors of the enzyme, withIC50s ranging from 360 to 1 
M. Adetailed study of product inhibition by the natural NS3 substrates isdescribed in the preceding paper[Steinkühler, C., et al. (1997) Biochemistry 37,8899-8905]. Here we report the results of a studyof the structure-activity relationship of the NS3 product inhibitors,which suggest that the mode of bindingof the P region-derived products is similar to the ground-state bindingof the corresponding substrates,with additional binding energy provided by the C-terminal carboxylate.Optimal binding requires a dualanchor: an "acid anchor" at the N terminus and a "P1 anchor"at the C-terminal part of the molecule.We have then optimized the sequence of the product inhibitors byusing single mutations and combinatorialpeptide libraries based on the most potent natural product,Ac-Asp-Glu-Met-Glu-Glu-Cys-OH (Ki =0.6M), derived from cleavage at the NS4A-NS4B junction. Bysequentially optimizing positions P2, P4,P3, and P5, we obtained several nanomolar inhibitors of the enzyme.These compounds are useful bothas a starting point for the development of peptidomimetic drugs and asstructural probes for investigatingthe substrate binding site of NS3 by modeling, NMR, andcrystallography.
M. Adetailed study of product inhibition by the natural NS3 substrates isdescribed in the preceding paper[Steinkühler, C., et al. (1997) Biochemistry 37,8899-8905]. Here we report the results of a studyof the structure-activity relationship of the NS3 product inhibitors,which suggest that the mode of bindingof the P region-derived products is similar to the ground-state bindingof the corresponding substrates,with additional binding energy provided by the C-terminal carboxylate.Optimal binding requires a dualanchor: an "acid anchor" at the N terminus and a "P1 anchor"at the C-terminal part of the molecule.We have then optimized the sequence of the product inhibitors byusing single mutations and combinatorialpeptide libraries based on the most potent natural product,Ac-Asp-Glu-Met-Glu-Glu-Cys-OH (Ki =0.6M), derived from cleavage at the NS4A-NS4B junction. Bysequentially optimizing positions P2, P4,P3, and P5, we obtained several nanomolar inhibitors of the enzyme.These compounds are useful bothas a starting point for the development of peptidomimetic drugs and asstructural probes for investigatingthe substrate binding site of NS3 by modeling, NMR, andcrystallography.