Strategy for purifying maltose binding protein fusion proteins by affinity precipitation
2008
Raghava, Smita | Aquil, Samina | Bhattacharyya, Sanchari | Varadarajan, Raghavan | Gupta, Munishwar N.
The maltose binding protein (MBP) affinity tag has been extensively used for protein purification. A commercial grade cationic starch could precipitate MBP or an MBP-tagged protein quantitatively by simultaneous addition of 10% (w/v) polyethylene glycol (PEG) and 50mM calcium chloride. The precipitated MBP or MBP-tagged protein could be selectively dissociated by suspending the precipitate in 1M NaCl. In the case of a soluble MBP fusion with a fragment of human immunodeficiency virus protein gp120, 38% of the contaminating proteins could be removed by precipitation with PEG/CaCl₂ and 100% of the fusion protein was recovered. In all cases, the purified proteins showed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the expected changes in fluorescence emission spectra upon binding to maltose.
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