(16) Assay Card

General Information
Summary MCoTI-II shown to possess no anti-microbial activity.
Condition Antibacterial activities were measured using a liquid-growth inhibition assay as described previously. Briefly, 10 uL from 2-fold serial dilutions of the peptides (100-0.2 uM, final concentrations) were incubated in 96-wells microtiter plates with a starting OD600 of 0.001. After a 24 h incubation at 25 C, the antibacterial activity was monitored by measuring the culture absorbance at 595 nm using a microplate reader. The antifungal activity against N. crassa used liquid-growth inhibition assay. Briefly, fungal spores (final concentration of 10-4 spores/mL) were suspended in a growth medium containing Potato Dextrose Broth [DIFCO, in half-strength, supplemented with tetracycline (10ug/mL) and cefotaxim (100ug/mL)], dispensed by aliquots of 90 uL into wells of a microplate containing 10 uL of the serial dilution of the peptides, and incubated for 48 h at 25 C in the dark. Growth of fungi was evaluated as above. Positive controls were obtained using insect antimicrobial peptides (thanatin and androctonin).
Result The peptides present no significant antimicrobial activity against microbial strains chosen for their very high sensitivity to antibiotics.
AssayType Anti-bacterial

Heitz A, Hernandez JF, Gagnon J, Hong TT, Pham TT, Nguyen TM, Le-Nguyen D, Chiche L (2001) Solution structure of the squash trypsin inhibitor MCoTI-II. A new family for cyclic knottins. Biochemistry 40:7973-83

Proteins Assayed MCoTI-II