Journal: Journal of Biological Chemistry

Article Title: Translocation of Group 1 Capsular Polysaccharide in Escherichia coli Serotype K30

doi: 10.1074/jbc.m308775200

Figure Lengend Snippet: FIG. 3. Purification of Wza and WzaHis6. The purification protocols are described under “Experimental Procedures,” and the figure shows a Coomassie Blue-stained SDS-PAGE gel of relevant fractions. Lanes 1–4 show the purification of Wza from SB 3–14-solubilized OM (lane 1) by sequential anion exchange chromatography (lane 2) and hydroxyapatite chromatography (lane 3). All three lanes show Wza migrating as a major band at 40 kDa (i.e. Wza monomer) when the samples are heated in SDS-sample buffer at 100 °C prior to electrophoresis. The purified Wza sample from lane 3 shows the multimeric form when the samples are run in the same buffer without heating (lane 4). Lanes 5 and 6 show heated and unheated samples of purified WzaHis6, respectively, confirm- ing that WzaHis6 also forms SDS-stable multimers.

Article Snippet: Fractions containing Wza were applied to a hydroxyapatite column (5-ml Econo-PacCHT-II, Bio-Rad) equilibrated with 50 mM sodium phosphate buffer, pH 7.0, containing 0.008% DDM and the protein was eluted with a linear gradient of 0.05–1 M sodium phosphate buffer, pH 7.0, containing 0.008% DDM.

Techniques: Purification, Staining, SDS Page, Chromatography, Electrophoresis