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Combining Metabolic Alkyne Labeling and Click Chemistry for Secretome Analysis of Serum‐Containing Conditioned Medium †
Author(s) -
Zheng Jiangnan,
Mao Yuan,
Feng Shun,
Tian Ruijun
Publication year - 2021
Publication title -
chinese journal of chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.28
H-Index - 41
eISSN - 1614-7065
pISSN - 1001-604X
DOI - 10.1002/cjoc.202000752
Subject(s) - bioorthogonal chemistry , alkyne , chemistry , click chemistry , azide , cysteine , cycloaddition , combinatorial chemistry , biochemistry , residue (chemistry) , organic chemistry , catalysis , enzyme
Main observation and conclusion Bioorthogonal click chemistry has emerged as a powerful tool for the specific modification of proteins in complex mixtures. Metabolic labeling of proteins with azide followed by the copper‐catalyzed azide−alkyne cycloaddition (CuAAC) with alkyne‐based affinity probes/beads is widely applied to study protein turnover and post‐translational modifications (PTMs). However, it has long been known that the alkyne‐based enrichment of high concentration protein samples ( e.g ., serum) is not sufficient to remove high abundant contaminating proteins. Herein, we demonstrate that the protein contamination is mainly caused by thiol‐yne addition between terminal alkyne and cysteine residue. Furthermore, we report that azide‐based enrichment combined with metabolic labeling with alkyne can significantly reduce contamination and improve the enrichment efficiency of secreted proteins in serum‐containing conditioned medium.