Fig. 1.

A, Growth curve profile of B. subtilis in minimal medium. A composite growth curve of B. subtilis grown in batch culture, in minimal medium under laboratory conditions. In each SILAC experiment cultures were labeled with the respective isotope of lysine and cells were harvested at the denoted time points. B, Phosphoproteomics workflow. Cells were harvested and lysed at the respective stages in each SILAC experiment. Proteins were extracted, mixed in equal ratios and digested with endoproteinase Lys-C. Digested peptides were fractionated by Offgel isoelectric focusing (for whole proteome analysis) and, separately, by SCX and TiO₂ chromatographies (for phosphopeptide enrichment). All samples were analyzed on LTQ-Orbitrap mass spectrometer.