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Quantitative and Comparative Profiling of Protease Substrates through a Genetically Encoded Multifunctional Photocrosslinker
Author(s) -
He Dan,
Xie Xiao,
Yang Fan,
Zhang Heng,
Su Haomiao,
Ge Yun,
Song Haiping,
Chen Peng R.
Publication year - 2017
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.201708151
Subject(s) - quantitative proteomics , proteomics , bioorthogonal chemistry , chemistry , proteolysis , protease , chaperone (clinical) , linker , biochemistry , computational biology , biology , combinatorial chemistry , click chemistry , enzyme , gene , medicine , pathology , computer science , operating system
A genetically encoded, multifunctional photocrosslinker was developed for quantitative and comparative proteomics. By bearing a bioorthogonal handle and a releasable linker in addition to its photoaffinity warhead, this probe enables the enrichment of transient and low‐abundance prey proteins after intracellular photocrosslinking and prey–bait separation, which can be subject to stable isotope dimethyl labeling and mass spectrometry analysis. This quantitative strategy (termed isoCAPP) allowed a comparative proteomic approach to be adopted to identify the proteolytic substrates of an E. coli protease–chaperone dual machinery DegP. Two newly identified substrates were subsequently confirmed by proteolysis experiments.