Abstract
NS4A is a non-structural multi-tasking small peptide that is essential for HCV maturation and replication. The central odd-numbered hydrophobic residues of NS4A (Val-23' to Leu-31')(i) are essential for activating NS3 upon NS3/4A protease complex formation. This study aims to design new specific allosteric NS3/4A protease inhibitors by mutating Val-23', Ile-25', and Ile-29' into bulkier amino acids. Pep-15, a synthetic peptide, showed higher binding affinity towards HCV-NS3 subtype-4 than native NS4A. The K-d of Pep-15 (80.0 +/- 8.0 nM) was twice as high as that of native NS4A (169 +/- 37 nM). The mutant Pep-15 inhibited the catalytic activity of HCV-NS3 by forming an inactive complex. Molecular dynamics simulations suggested that a cascade of conformational changes occurred, especially in the catalytic triad arrangements, thereby inactivating NS3. A large shift in the position of Ser-139 was observed, leading to loss of critical hydrogen bonding with His-57. Even though this study is not a classic drug discovery study-nor do we propose Pep-15 as a drug candidate-it serves as a stepping stone towards developing a potent inhibitor of hitherto untargeted HCV subtypes. (C) 2020 The Authors. Published by Elsevier B.V. on behalf of Cairo University. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).