In addition, the non-substrate based inhibitors, such as small molecule inhibitors, showed significant inhibitory activities at low micromolar concentrations against the flavivirus proteases [31, 32]. Although several of these compounds are potent inhibitors of the dengue NS2b-NS3

protease, some showed poor stability in solution. Furthermore, several studies did not use Cell Cycle inhibitor cell-based assays to evaluate the toxicity and antiviral efficacy of the identified compounds [18]. The nature of the dengue protease, which possesses a flat and hydrophilic active site, decreases the possibility of identifying potent inhibitors to develop as antiviral therapeutics [18]. Based on the results of this buy CHIR98014 study, we postulate that the hydrophobic residues of Ltc 1 are important for stabilising the binding to the hydrophilic active site of the dengue protease. In this study, the inhibitory potential of the Ltc 1 peptide against the dengue protease was further verified using cell based assays. Previously, other characteristics of the latarcin family peptides, such as anti-neoplastic cells activities [21], were examined. The latarcin peptides can alter the lipid bilayers of the cell membrane, may induce the apoptosis of mammalian cells [21]. Because of this, the possible effect of the Ltc 1 peptide on cell proliferation was removed to avoid false interpretation

of AZD2281 the antiviral activity. Subsequently, the antiviral activity of the Ltc 1 peptide was evaluated at the doses with minimal effects on cell proliferation as determined by MTT assay and Real-Time Cellular Analysis (RTCA). The results of the immunostaining and western blot analyses showed that the Ltc 1 peptide significantly reduced the viral particles and non-structural protein NS1 in DENV-infected cells. Furthermore, the results of the time-of-addition assay showed that the Ltc 1 peptide inhibited dengue virus replication at both the simultaneous and post-treatments compared to the pre-treatment. The mechanism of antimicrobial activity of the latarcin peptides depends on the helix-hinge-helix structure that is important for lysing

bacterial cell membranes [35, 36]. This finding emphasised that the direct incubation of DENV with the Ltc 1 peptide during Rucaparib molecular weight the simultaneous treatment may led to lysis of the viral particles by the peptide. The results of the post-treatment and dose-response assays showed that the viral load was significantly deceased after treatment with the Ltc 1 peptide. Based on this finding, we hypothesise that the Ltc 1 peptide may interrupt the dengue life cycle in HepG2 cells during post-translational processing of the polyprotein by inhibiting the dengue serine protease. This inhibition may hinder flavivirus replication and virion assembly, as evidenced by the lack of infectious virion production in mutants carrying inactivating viral proteases [13].