Degradative processes within the lysosomal system are crucial mechanisms by which homeostasis is maintained in the cell. One family of proteases that are shown to be optimally active within the lysosome’s acidic environment are the cysteine cathepsins. Initially, these cathepsins were believed to have non-specific functions in the endolysosomal compartment, simply breaking down the unwanted cellular proteins. However, recent research suggests both distinct as well as redundant functions for individual cathepsins. Importantly, aberrant cathepsin functions are implicated in numerous human diseases. Two prominent members of the family are cathepsin L and B enzymes that play critically important roles in both normal physiology and diseased states. Despite this, functional understanding of these enzymes in appropriate cellular contexts are not well understood. In this work, we report the development of highly selective and clickable probes of cathepsins L and B enzymes. The design of the probes exploits the key differences between the catalytic activities of the two enzymes: cathepsin L engages mainly in endopeptidase activity, while cathepsin B partakes in both endopeptidase and exopeptidase processes. The developed probes are highly selective, cell permeable, and particularly sensitive to the activity states of the two cathepsins. We anticipate that the probes will find extensive applications in dissecting the cellular roles of both cathepsins at the proteome levels.

Selective activity-based, tagless probes of human cathepsin L and B enzymes for functional proteomics

ACS Spring Meeting 2021

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