Quantitative and Comparative Profiling of Protease Substrates through a Genetically Encoded Multifunctional Photocrosslinker
Dan He
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
These authors contributed equally.
Search for more papers by this authorXiao Xie
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
These authors contributed equally.
Search for more papers by this authorFan Yang
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHeng Zhang
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHaomiao Su
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, 430072 China
Search for more papers by this authorYun Ge
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHaiping Song
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorCorresponding Author
Prof. Peng R. Chen
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, 100871 China
Search for more papers by this authorDan He
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
These authors contributed equally.
Search for more papers by this authorXiao Xie
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
These authors contributed equally.
Search for more papers by this authorFan Yang
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHeng Zhang
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHaomiao Su
College of Chemistry and Molecular Sciences, Wuhan University, Wuhan, 430072 China
Search for more papers by this authorYun Ge
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorHaiping Song
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Search for more papers by this authorCorresponding Author
Prof. Peng R. Chen
Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing, 100871 China
Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, 100871 China
Search for more papers by this authorGraphical Abstract
A genetically encoded trifunctional photocrosslinker was developed that bears a bioorthogonal handle and a releasable linker in addition to its photoaffinity warhead. This probe enabled the enrichment of transient and low-abundance prey proteins after intracellular photocrosslinking and prey–bait separation. A quantitative and comparative proteomic strategy using this probe revealed the substrates of a protease.
Abstract
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.
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Citing Literature
November 13, 2017
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