Characterization and kinetics of a cathepsin B-inhibiting protein from Musa acuminata Colla peel.

Hyperexpression of cathepsin B caused by an imbalance of endogenous inhibitors is involved in multiple pathologies, hence making it a key therapeutic target. Protease inhibitors are effective biomolecules that regulate protease activities and are considered potential therapeutic agents in various diseases. Plant protease inhibitors have been reported as an effective complementary alternative drug. A proteinaceous cathepsin B inhibitor (CBI-BP) has been isolated from Musa acuminata Colla (banana) peel with a molecular weight of 27.9 kDa on SDS-PAGE. The purity of the CBI-BP was confirmed on the native- PAGE. The isolated CBI-BP showed an IC₅₀ value of 8.14 μg and a K_i value of 10.59 μg (0.19 μM). Cathepsin B inhibition kinetics indicated that CBI-BP follows a mixed-type of cathepsin B inhibition. Its inhibition activity was also confirmed by reverse zymography. The inhibitor was stable from pH 2.6-10.0 with maximum activity at pH 7.2, temperature 25-100 °C and exhibited thermostability for 60 min at 70 °C. MALDI/TOF/MS analysis of CBI-BP showed 40 % similarity to the GH18 domain-containing protein (A0A4S8JRM9) from Musa balbisiana. Although in-silico docking studies showed binding of A0A4S8JRM9 to cathepsin B affects the binding energy of the substrate to cathepsin B but is not reported for any anti-cathepsin B activity. This suggests that isolated CBI-BP might be a novel protein with anti-cathepsin B activity. Thus the isolated CBI-BP may be further explored as possible anti-cathepsin B drug.