Whole-cell proteins were obtained from the S. Typhimurium strain SH100, a derivative of ATCC 14028, with the stringent response induced by serine hydroxamate, as described previously [26]. Agarose 2-DE was performed at least three times on independent samples. More than 350 protein spots from the strain were detected on each 2-DE gel stained with Coomassie Brilliant Blue. To identify proteins on the agarose 2-DE gels, we excised 230 spots from the 12% gel and 136 spots from the 15% gel. We finally identified

RG7112 a total of 360 proteins (273 proteins from the 12% gel [Figure 1A] and 87 proteins from the 15% gel [Figure 1B]) by MS/MS analysis out of 307 protein spots (232 spots from the 12% gel and 75 spots from the 15% gel) that were successfully excised (see additional file: 1). In total, 267 proteins were obtained from the gels, with 40 proteins identified as being redundant. The highest and lowest molecular masses of identified proteins were 93.4 kDa for AcnB (aconitate hydrase 2, spot 188) and 7.4 kDa for CspC (cold-shock protein, spot 303), respectively. Fifty spots (35 spots from the 12% gel and 15 spots from the 15% gel) were found in a basic range.

Interestingly, 78 protein spots (25.4%) were annotated as putative proteins on the genome of the S. Typhimurium LT2 strain, which is more than 98% identical in sequence to the 14028 strain [27]. Figure 1 Agarose 2-DE reference map of the S . Typhimurium strain SH100, prepared using a 12% gel focused on high-molecular-mass proteins (A) and a 15% gel focused on low-molecular-mass Vistusertib chemical structure proteins (B). Strain SH100 was grown under amino acid starvation as described previously [26]. Gels are stained with Coomassie Brilliant Blue. Identified spots are numbered (corresponding to the spot numbers Methane monooxygenase in additional file: 1. Proteins identified on the reference map). We estimated the molecular weight of the protein spots on the 2-DE gels and compared them with the theoretical molecular weight of strain SH100. While most of the estimated molecular weight values matched the theoretical values,

we found 14 protein spots on the map that had different experimental and predicted molecular weights values (Figure 2). These proteins might be post-translationally modified by proteolytic processing, phosporylatoin of multiple amino acid residues and/or an artifact caused by Erismodegib sample preparation. For example, the experimental molecular weight of OmpA indicated that the protein was likely processed by a proteolytic enzyme, because two different spots (spot nos. 152 and 287) were identified as OmpA, the experimental masses of which were significantly lower than the theoretical values. Similar results were described in other reports [28, 29]. Figure 2 Comparison of the gel-estimated and theoretically calculated molecular weight (Mw) of the identified protein spots.