I want to digest sumo/His label when the protein in still hang on the Ni column. I wash the undesired proteins as usual, then balance the column with 3ml enzyme digestion buffer, then add sumo enzyme for digesting the fusion protein at 4℃ for 12h in 4ml sumo digesting buffer. Then catch the flow through liquid, take some sample from flow through liquid and beads after elution for SDS-PAGE, the brand explains the label did not have been cut, it is still on the beads entirely.
How do you purify SUMO tagged proteins?
Sumo protease cleavage and clean up Change protein buffer to Sumo cleavage buffer. Either dialyze O/N or concentrate and dilute protein in cleavage buffer. If protein becomes cloudy, spin in tabletop centrifuge 4100 rpm, 15 min. Add 30 microliters SUMO protease (0.148 mg/ml; current prep) per liter of starting culture.
Sumo protease cleavage and clean up Change protein buffer to Sumo cleavage buffer. Either dialyze O/N or concentrate and dilute protein in cleavage buffer. If protein becomes cloudy, spin in tabletop centrifuge 4100 rpm, 15 min. Add 30 microliters SUMO protease (0.148 mg/ml; current prep) per liter of starting culture.
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