BioMed Research International / 2014 / Article / Fig 9

Research Article

C-Terminal Domain Swapping of SSB Changes the Size of the ssDNA Binding Site

Figure 9

Gel-filtration chromatographic analyses of KpSSBnStSSBc and KpSSBnPaSSBc. Purified protein (2 mg/mL) was applied to a Superdex 200 HR 10/30 column (GE Healthcare Bio-Sciences, Piscataway, NJ, USA) equilibrated with Buffer D. The column was operated at a flow rate of 0.5 mL/min, and 0.5 mL fractions were collected. The proteins were detected by measuring the absorbance at 280 nm. The column was calibrated with proteins of known molecular weight: thyroglobulin (670 kDa), γ-globulin (158 kDa), ovalbumin (44 kDa), and myoglobin (17 kDa). The values for the standard proteins and the SSB variants were calculated from the equation: , where is column void volume, is elution volume, and is geometric column volume.