A Hexylchloride-Based Catch-and-Release System for Chemical Proteomic Applications

Bioorthogonal ligation methods that
allow the selective conjugation
of fluorophores or biotin to proteins and small molecule probes that
contain inert chemical handles are an important component of many
chemical proteomic strategies. Here, we present a new catch-and-release
enrichment strategy that utilizes a hexylchloride group as a bioorthogonal
chemical handle. Proteins and small molecules that contain a hexylchloride
tag can be efficiently captured by an immobilized version of the self-labeling
protein HaloTag. Furthermore, by using a HaloTag fusion protein that
contains a protease cleavage site, captured proteins can be selectively
eluted under mild conditions. We demonstrate the utility of the hexylchloride-based
catch-and-release strategy by enriching protein kinases that are covalently
and noncovalently bound to ATP-binding site-directed probes from mammalian
cell lysates. Our catch-and-release system creates new possibilities
for profiling enzyme families and for the identification of the cellular
targets of bioactive small molecules.