Journal
JOURNAL OF PROTEOME RESEARCH
Volume 12, Issue 12, Pages 5978-5988Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr400775w
Keywords
quantitative proteomics; mass spectrometry; Escherichia coli; SILACE; IPTG induction; QconCAT compatible labeling method
Categories
Funding
- National Basic Research Programs [2011CB910600, 2013CB911201]
- National Natural Science Foundation of China [31070673, 31170780]
- National High-Tech Research and Development Program of China
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Stable isotope labeling by amino acids in cell culture (SILAC) has been widely used in yeast, mammalian cells, and even some multicellular organisms. However, the lack of optimized SILAC media limits its application in Escherichia coli, the most commonly used model organism. We optimized SILACE medium (SILAC medium created in this study for E. coli) for nonauxotrophic E. coli with high growth speed and complete labeling efficiency of the whole proteome in 12 generations. We applied a swapped SILAC workflow and pure null experiment with the SILACE medium using E. coli BL21 (DE3) cells hosting a recombinant plasmid coding for glutathione-S-transferase 05 (GST) and ubiquitin binding domain before and after isopropyl thiogalactoside (IPTG) induction. Finally, we identified 1251 proteins with a significant change in abundance. Pathway analysis suggested that cell growth and fissiparism were inhibited accompanied by the down-regulation of proteins related to energy and metabolism, cell division, and the cell cycle, resulting in the size and shape change of the induced cells. Taken together, the results confirm the development of SILACE medium suitable for efficient and complete labeling of E. coli cells and a data filtering strategy for SILAC-based quantitative proteomics studies of E. coli.
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