Inhibition of Recombinant Human Cathepsin K by Natural and Synthetic Inhibitors

Cathepsin K, a lysosomal cysteine protease, is abundantly expressed in osteoclast cells. The enzyme has been considered a target for osteoporosis therapy. The objective of the research was to prepare functional recombinant cathepsin K and used it in the development of new cathepsin K inhibitors. We have previously overexpressed human procathepsin K in E. coli and activated the enzyme with the aid of porcine pepsin. The cathepsin K activity was measured at pH 5.5 using chromogenic substrate, Z‐FR‐pNA. The activity was monitored by observing the increase in absorbance at 405 nm caused by the release of pNA. The activated cathepsin K has been tested against two groups of inhibitors: natural and synthetic (provided by the synthesis group). The natural inhibitors consisted of several flavonoids. Apigenin, myricetin, and celastrol are among the stronger inhibitors. The synthetic inhibitors consisted of eight different tripeptide analogs. Their IC50 values and kinetic data were compared. Supported by National Cancer Institute at NIH (Grant No. 3R15CA086933‐04 and 3R15CA086933‐04A2S1) and Western Illinois University.