Design of selective eglin inhibitors of HCV NS3 proteinase.
Hepatitis C virus (HCV) infection is a major health problem that leads to cirrhosis and hepatocellular carcinoma in a substantial number of infected individuals, estimated to be 100-200 million worldwide. Unfortunately, immunotherapy or other effective treatments for HCV infection are not yet available, and interferon administration has limited efficacy. Different approaches to HCV therapy are being explored, and these include inhibition of the viral proteinase, helicase, and RNA-dependent RNA polymerase and development of a vaccine. Here we present the design of selective inhibitors with nanomolar potencies of HCV NS3 proteinase based on eglin c. These eglin c mutants were generated by reshaping the inhibitor active site-binding loop, and the results emphasize the role played by residues P5-P4' in enzyme recognition. In addition, alanine scanning experiments provide evidence that the N terminus of eglin c also contributes to NS3 binding. These eglin inhibitors offer a unique tool for accurately assessing the requirements for effective inhibition of the enzymatic activity of NS3 and at the same time can be considered lead compounds for the identification of other NS3 inhibitors in targeted design efforts.
Department of Protein Engineering & Biocrystallography, Istituto di Ricerche di Biologia Molecolare (IRBM) P. Angeletti, Roma, Italy.
F
F
Martin
N
N
Dimasi
C
C
Volpari
C
C
Perrera
S
S
Di Marco
M
M
Brunetti
C
C
Steinkühler
R
R
De Francesco
M
M
Sollazzo
D000818:Animals; D016174:Hepacivirus / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D007865:Leeches; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D015202:Protein Engineering / Q000379:methods; D011506:Proteins; D020365:RNA Helicases; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology; D015843:Serpins / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013816:Thermodynamics; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors*; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D046911:Macromolecular Substances; D010446:Peptide Fragments; D011506:Proteins; D020365:RNA Helicases; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors; D015843:Serpins; D017361:Viral Nonstructural Proteins; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C028656:eglin proteinase inhibitors