Boston Biochem in March 2018 Edition of Nature Communications

Reactive-site-centric chemoproteomics identifies a distinct class of deubiquitinase enzymes

Close on the heels of last October’s Nature publication “USP7 Small Molecule Inhibitors Interfere with Ubiquitin Binding,” Bio-Techne has a second publication, this time in Nature Communications.  Published March 21 online, the article “Reactive-site-centric chemoproteomics identifies a distinct class of deubiquitinase enzymes” marks a second collaborative effort between Genentech scientists and Boston Biochem (Bio-Techne, Cambridge), this time with contributions from scientists at Stanford University and UbiQ Bio.

 

The article describes the development and characterization of new Activity-Based Probe (ABP) variants based on Ubiquitin and Ubiquitin-like proteins conjugated to a vinyl sulfone “warhead.”  ABPs are powerful tools to study enzymatic activity in complex biological systems, and are well suited to the study of enzymes possessing nucleophilic active-site residues, including deubiquitinating enzymes (DUBs).   ABPs can be used to monitor the activity of many DUBs simultaneously, providing a snapshot of the active enzymes in various biological conditions.  However, inferring DUB activity using ABPs relies on the assumptions that the probe and enzyme have undergone a covalent reaction specifically at the active site, and that inactive enzymes will not react with the probe.  Testing these assumptions is challenging and requires protein-by-protein validation.  To address such limitations, we modified the design of a Ubiquitin-based ABP and optimized enrichment methodologies to permit the detection of probe-labeled residues by mass spectrometry (MS).  This approach confirmed the expected labeling of DUB catalytic cysteine residues by ABPs, but also revealed unexpected widespread labeling of non-catalytic cysteine residues across DUB enzymes and of non-DUB proteins.  Using this method, we identified a previously unannotated DUB (ZUFSP) that represents a unique class of DUB enzymes.

 

Boston Biochem scientists Dr. Carsten Schwerdtfeger and Dr. Greg Costakes made significant contributions to the publication, and Boston Biochem will offer the new activity probes commercially in the very near future.

 

Read the Full Nature Communications Article Here - https://www.nature.com/articles/s41467-018-03511-6.pdf