Protein acetylation in response to carbon overflow: detecting dynamic acetylation changes by mass spectrometry
Birgit Schilling, David Christensen, Bradford W. Gibson, Alan J. Wolfe et al.
Experimental Design for the E. coli glucose dependent acetylome project:
To identify specific lysines with significant acetylation changes, we grew cells in TB7 with and without glucose, applied label-free, quantitative mass spectrometry (MS1 Filtering), monitored acetylated peptides, and compared acetylation changes (i) over time during growth in the presence of glucose and (ii) between glucose and no glucose growth conditions. This approach revealed increases over time in both the number of detected lysine acetylation sites and the dynamic relative increase of determined acetylation levels. During the time course, the change in acetylation varied greatly between individual lysines from the same proteins, consistent with differences in specificity. Glucose-regulated lysine acetylation sites were particularly predominant in central metabolic pathways and overlapped extensively with acetylPhosphgate-regulated lysine acetylation sites.
MS/MS spectral library for all identified E. coli 2813 lysine acetylation sites from 780 acetylated proteins (see above imported as Targeted MS Runs)
Quantification of glucose regulated acetylation – dynamic acetylation changes following a glucose time course displayed for >400 proteins (download graphical displays).
Legend: Differential rates of acetylation: Individual Kac sites from the same protein show different rates of increasing acetylation during the glucose time course experiments (all Kac fold changes were normalized for potential protein level changes). Dynamic change of acetylation is displayed for >400 E coli proteins with the following Kac fold changes monitored between the following Glc-supplemented time points, 2h/2h (fold change set to 1), 5h/2h, 8h/2h, and 12h/2h.
MS/MS identification details for lysine acetylated peptides from database search results, Protein Pilot and Mascot (download identification excel file)
3 dimensional protein structures (Pymol) of selected acetylated proteins from the TCA cycle, Glycolysis and Phosphate Pentose Pathway indicating location of acetylation (Kac) sites (download pdf file)