Molecular Characterization and Cytotoxic Activity of McTI, a Novel Cystine-Knot Inhibitor from Momordica charantia.

Abstract

Medicinal plants are rich source of pharmaceutically active peptides and proteins. Momordica charantia, a traditional medicinal plant commonly known as bitter melon, is a member of a family Cucurbitaceae and has been explored for various human diseases such as diabetes, peptic ulcer, malaria, infectious diseases, and cancer. Yet, to date only a handful of studies are available related to the therapeutic potential of Momordica charantia seed proteins. In present study, a novel trypsin inhibitor McTI was purified by 2D-LC technique from ammonium sulphate precipitated crude seed proteins of Momordica charantia. The crude seed proteins and gel filtration fractions were explored for their cytotoxic response against MDA-MB-231 cells. Later, for deeper understanding, complete amino acid sequence of McTI was established by Edman protein sequencing and 3D structure was predicted by comparative homology modeling using template trypsin inhibitor 3 from spiny bitter cucumber. In silico molecular docking and dynamic simulation experiments were also performed to study the interaction of McTI with bovine trypsin. The results revealed that McTI is a 30 amino acids peptide having a molecular mass of 3388.4 Da and showed sequence similarity with previously reported cystine knot trypsin inhibitors from plant. McTI is a disulfide rich peptide having 6 cysteine residues that can form 3 disulfide bonds (Cys3-Cys20, Cys10-Cys22 and Cys16-Cys28). Six hydrogen bond interactions of McTI with bovine trypsin were observed in molecular docking whereas, additional hydrogen bond interactions were noticed in molecular dynamic simulation studies. In cytotoxicity analysis against MDA-MB-231 cells, crude seed proteins and F12 exhibited a significant dose dependent response with an IC 50 values of 82.10 ± 6.46 µg/ml and 81.13 ± 4.26, respectively. These findings suggest Momordica charantia proteins as a possible anticancer agent. Furthermore, McTI is a valuable addition in the squash inhibitor family.