In situ activity-based protein profiling of serine hydrolases in E. coli

Publication date: September 2014 Source:EuPA Open Proteomics, Volume 4 Author(s): Dmitry Shamshurin , Oleg V. Krokhin , David Levin , Richard Sparling , John A. Wilkins A fluorophosphonate based alkyne activity probe was used for the selective labeling of active serine hydrolases in intact Escherichia coli cells. A biotin-azide tag was subsequently attached to the alkyne functionality of the probe with copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction. Comparison of proteins from in-cell and lysate labeled preparations suggested qualitatively similar patterns of reactivity in both preparations. Approximately 68%, 30 of the total 44 serine hydrolases detectable in E. coli were labeled with the probe indicating significant coverage with a single probe. The methods described here offer a useful tool for profiling and monitoring serine hydrolase activity in situ. Graphical abstract
Source: EuPA Open Proteomics - Category: Bioinformatics Source Type: research