Table 1 Comparison of the purification of extra- and intracellular CGTases by starch adsorption and by affinity chromatography with Ni-Sepharose 6
Fraction | Volume (ml) | Protein conc. (mg/ml) | Total protein (mg) | Volume activity (U/ml) | Total activity (U) | Specific activity (U/mg) | Yield (%) | Purification (-fold) |
|---|---|---|---|---|---|---|---|---|
(a) Purification of the intracellular PelB-CGTase by batch wise starch adsorption | ||||||||
Crude extract | 15 | 24.7 | 370.5 | 330 | 4950 | 13.4 | 100 | 1.0 |
Washing step | 30 | 2.1 | 63.9 | 37.5 | 1123.8 | 17.6 | 22.7 | 1.3 |
Eluate | 2 | 0.22 | 0.43 | 324 | 648 | 1505.0 | 13.1 | 112.6 |
(b) Purification of the extracellular DacD-CGTase by batch wise starch adsorption | ||||||||
Crude extract | 100 | 0.079 | 7.94 | 14.4 | 1436.9 | 181.1 | 100 | 1.0 |
Washing step | 90 | 0.006 | 0.58 | 2.7 | 239.3 | 411.7 | 16.7 | 2.3 |
Eluate | 1.5 | 0.272 | 0.41 | 445.4 | 668.0 | 1636.9 | 46.5 | 9.0 |
(c) Purification of the extracellular DacD-CGTase by affinity chromatography with Ni-Sepharose 6 | ||||||||
Crude extract | 100 | 0.079 | 7.94 | 14.4 | 1436.9 | 181.1 | 100 | 1.0 |
Unbound | 100 | 0.072 | 7.20 | 9.9 | 988.8 | 137.4 | 68.8 | 0.8 |
Washing step | 2.5 | 0.043 | 0.11 | 13.5 | 33.8 | 313.3 | 2.4 | 1.7 |
Eluate | 3.5 | 0.020 | 0.07 | 11.7 | 41.0 | 595.6 | 2.9 | 3.3 |