TY - JOUR
T1 - Discovery of Electrophiles and Profiling of Enzyme Cofactors
AU - Dettling, Suzanne E.
AU - Ahmadi, Mina
AU - Lin, Zongtao
AU - He, Lin
AU - Matthews, Megan L.
N1 - Funding Information:
This work was supported by the University of Pennsylvania (M.L.M) and an NIH DP1 grant (no. 1DP1DA051620). The authors would like to thank Radu M. Suciu (Department of Chemistry, The Scripps Research Institute) and Hee Jong Kim (Biochemistry & Molecular Biophysics Graduate Group, University of Pennsylvania) for uploading the CIMAGE script to GitHub.
Funding Information:
This work was supported by the University of Pennsylvania (M.L.M) and an NIH DP1 grant (no. 1DP1DA051620). The authors would like to thank Radu M. Suciu (Department of Chemistry, The Scripps Research Institute) and Hee Jong Kim (Biochemistry & Molecular Biophysics Graduate Group, University of Pennsylvania) for uploading the CIMAGE script to GitHub.
Publisher Copyright:
© 2020 The Authors.
PY - 2020/12
Y1 - 2020/12
N2 - Reverse-polarity activity-based protein profiling (RP-ABPP) is a chemical proteomics approach that uses nucleophilic probes amenable to “click” chemistry deployed into living cells in culture to capture, immunoprecipitate, and identify protein-bound electrophiles. RP-ABPP is used to characterize the structure and function of reactive electrophilic post-translational modifications (PTMs) and the proteins harboring them, which may uncover unknown or novel functions. RP-ABPP has demonstrated utility as a versatile method to monitor the metabolic regulation of electrophilic cofactors, using a pyruvoyl cofactor in S-adenosyl-L-methionine decarboxylase (AMD1), and to discover novel types of electrophilic modifications on proteins in human cells, such as the glyoxylyl modification on secernin-3 (SCRN3). These cofactors cannot be predicted by sequence, and therefore this area is relatively undeveloped. RP-ABPP is the only global, unbiased approach to discover such electrophiles. Here, we describe the utility of these experiments and provide a detailed protocol for de novo discovery, quantitation, and global profiling of electrophilic functionality of proteins.
AB - Reverse-polarity activity-based protein profiling (RP-ABPP) is a chemical proteomics approach that uses nucleophilic probes amenable to “click” chemistry deployed into living cells in culture to capture, immunoprecipitate, and identify protein-bound electrophiles. RP-ABPP is used to characterize the structure and function of reactive electrophilic post-translational modifications (PTMs) and the proteins harboring them, which may uncover unknown or novel functions. RP-ABPP has demonstrated utility as a versatile method to monitor the metabolic regulation of electrophilic cofactors, using a pyruvoyl cofactor in S-adenosyl-L-methionine decarboxylase (AMD1), and to discover novel types of electrophilic modifications on proteins in human cells, such as the glyoxylyl modification on secernin-3 (SCRN3). These cofactors cannot be predicted by sequence, and therefore this area is relatively undeveloped. RP-ABPP is the only global, unbiased approach to discover such electrophiles. Here, we describe the utility of these experiments and provide a detailed protocol for de novo discovery, quantitation, and global profiling of electrophilic functionality of proteins.
KW - chemoproteomics
KW - de novo PTM discovery
KW - enzyme activity
KW - post-translational modifications
KW - reverse-polarity activity-based protein profiling
UR - http://www.scopus.com/inward/record.url?scp=85096245683&partnerID=8YFLogxK
U2 - 10.1002/cpch.86
DO - 10.1002/cpch.86
M3 - Article
C2 - 33197155
AN - SCOPUS:85096245683
SN - 2160-4762
VL - 12
JO - Current protocols in chemical biology
JF - Current protocols in chemical biology
IS - 4
M1 - e86
ER -