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A nascent proteome study combining click chemistry with 2 DE
Author(s) -
Osterman Ilya A.,
Ustinov Alexey V.,
Evdokimov Denis V.,
Korshun Vladimir A.,
Sergiev Petr V.,
Serebryakova Marina V.,
Demina Irina A.,
Galyamina Maria A.,
Govorun Vadim M.,
Dontsova Olga A.
Publication year - 2013
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201200393
Subject(s) - proteome , click chemistry , chemistry , methionine , protein biosynthesis , proteomics , biochemistry , erythromycin , combinatorial chemistry , antibiotics , amino acid , gene
To investigate the dynamic cellular response to a condition change, selective labeling of the nascent proteome is necessary. Here, we report a method combining click chemistry protein labeling with 2D DIGE . To test the relevance of the method, we compared nascent proteomes of actively growing bacterial cells with that of cells exposed to protein synthesis inhibitor, erythromycin. Cells were incubated with methionine analog, homopropargyl glycin, and their nascent proteome was selectively labeled with monosulfonated neutral C y3 and C y5 azides specially synthesized for this purpose. Following fluorescent labeling, the protein samples were mixed and subjected to standard 2D DIGE separation. The method allowed us to reveal a dramatic reduction of newly synthesized proteins upon erythromycin treatment, while the total proteome was not significantly affected. Additionally, several proteins, whose synthesis was resistant to erythromycin, were identified.