Reductive activation markedly increases the stability of cathepsins B and L to extracellular ionic conditions.

Cathepsins B and L are thought to function extracellularly in pathological conditions. pH-Activity profiles of cathepsin B, measured in phosphate and acetate-Mes-Tris buffers of constant ionic strength, indicated that cathepsin B is sensitive to specific buffer ions, as previously reported for cathepsin L. In assessing the activity of these enzymes in vitro the influence of the buffer must therefore be taken into account. In Hank's balanced salt solution, a buffer modeling the extracellular fluid, the half-life of activated human liver cathepsin B at 37 degrees C is 245 +/- 11.3 s, at pH 7.2, and 857 +/- 50.1 s, at pH 6.8 (the peritumor pH), indicating that cathepsin B is markedly stable under these conditions. The stability was increased by the additional presence of proteins. Without immediate activation, however, the stabilities of both cathepsins B and L were markedly decreased, a large proportion of their activity being lost before it could be measured. Enzymes injected into the extracellular space in the unactivated state would therefore survive for only a very short time in their native conformation. It is proposed that the active site thiolate-imidazolium ion pair contributes substantially to the stability of cathepsins B and L to extracellular ionic conditions.