The Inhibition of Cathepsin S by its Propeptide — Specificity and Mechanism of Action

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Abstract
The interaction of human recombinant full‐length cathepsin S propeptide (amino acids 16–114) with mature cysteine proteinases was studied with respect to selectivity and pH dependence. The inhibitory capacity was tested towards mature human recombinant cathepsin S, purified cathepsin L from rat and Paramecium tetraurelia, rat cathepsin B, human cathepsin H, and papain. The propeptide of cathepsin S strongly inhibited cathepsin S (Ki= 0.27 nM) and the two cathepsin L species (Kt= 0.36 nM) at neutral pH. Papain, and to a minor extent cathepsin H, hydrolyzed the propeptide of cathepsin S, leading to competition with the hydrolysis of the fluorogenic substrates in the respective assays. Cathepsin B activity was nearly unaffected up to micromolar propeptide concentrations in the assay. The inhibition of cath‐epsin‐L‐like peptidases was diminished with decreasing pH, probably due to dramatic changes in the conformation of the propeptide. This assumption was supported by far‐ultraviolet CD spectroscopy and by the finding of rapid hydrolysis of the cathepsin S propeptide by cathepsin L at pH values less than 5.5.