Kinetically-controlled mechanism-based isolation of metabolic serine hydrolases in active form from complex proteomes: butyrylcholinesterase as a case study.

ABSTRACT: In this work an activity-based probe containing a carbamate group was designed to isolate human butyrylcholinesterase (hBChE), a metabolic serine hydrolase (mSH), from complex proteomes. The method took advantage of the native interaction mechanism of mSHs with carbamate pseudo-substrates for temporarily capturing the enzyme on a resin functionalized with the carbamate probe and releasing the enzyme in active form after removal of the contaminating proteins. The isolation relied on the possibility of manipulating the carbamylation and decarbamylation kinetics favoring the former during the capture and wash steps and the latter in the release step. The designed probe captured and released all the active hBChE isoenzymes present in plasma with high selectivity (up to ∼2000-fold purification)

SUBMITTER: Liu H 

PROVIDER: S-EPMC9075836 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature