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The interrogation of enzyme activity involves the ensemble averaging of many cells, loss of spatial relationships and is often biased to abundant proteins. Here the authors develop activity-dependent proximity ligation to quantify enzyme activity at the cellular and sub-cellular level in relevant biological contexts.

The Ras-family small GTPase RAB25 can exert both pro- and anti-oncogenic functions. Here, the authors develop all-hydrocarbon stabilized peptides targeting RAB25 and influencing the context-specificity phenotypes in cancer cell lines.