| Bioactivity |
SPECFIC ACTIVITY: After auto-catalytical activation at pH 4.0 the specific activity is >1000 mU/mg (international unit 1µmole/min/mg). ACTIVATION: Procathepsin K can be activated by adjusting the pH 4.0 by adding an equal volume of 100mM NaAc pH 3.9, 10 mM DTT, 5 mM EDTA followed by incubation for 40 min at RT. Activated mature Cathepsin is highly auto-proteolytic at pH 4.0, and care must be taken to avoid self-proteolysis. If the activated enzyme is not used immediately, we recommend to add methyl methanthiosulfonate (1 mM final concentration; MeS-SO2Me; MMTS) and to freeze the sample in liquid nitrogen or on dry ice. The hydrophobic thiol-reactive compound MMTS modifies cysteine’s by attaching its relatively small, uncharged thiomethyl-blocking group to reactive sulfhydryl groups. This reversible reaction arrests the autoproteolytic process. The activity of the enzyme can be restored to nearly unmodified levels by adding L-cysteine (3M excess over MMTS) to the enzyme solution.
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SDS-PAGE showing ab123765.