FIGURE 9.

Bioactivities of cyclotides. A, hemolytic dose-response curves were obtained using 2-fold dilutions starting with 50 μm peptide. Peptide solutions were tested in triplicate with a 0.25% (v/v) suspension of human RBCs and incubated for 1 h at 37 or at 4 °C. The peptide concentration needed to cause 50% of hemolysis (HC₅₀) was determined by fitting a sigmoidal dose-response curve. Melittin was included as a positive control. None of the cyclotides tested induced hemolysis at 4 °C. B, activity of cyclotides against E. coli. The survival percentage is plotted as function of peptide concentration. For clarity, only kB1, kB2, and cyO2 are shown. The errors bars represent the S.E. None of the cyclotides tested showed antimicrobial activity except cyO2; the MIC₅₀ for cyO2 as determined with a nonlinear regression analysis is 6.8 ± 0.4 μm. C, HIV virucidal activity of cyclotides. The ability of kB1, kB2, and cyO2 to inactivate HIV-1 NL4.3, or HIV-1 Clade A is shown. The errors bars represent the S.E. The concentration of cyclotide needed to cause a 50% reduction in viral infectivity (VC₅₀), compared with untreated virus, was calculated by a nonlinear regression analysis and compared with curves previously obtained for kB1 (21).