MBP-fusion protein purification problems

MaikelG me at work.anywhere
Fri Nov 19 05:34:41 EST 2004


Dear all,

For quite some time we are experiencing problems using the pMAL protein 
fusion and purification system of New England Biolabs. 

We produce our fusion protein in Sf9 cells, the protein is there (right 
size appearing on Western blot probed with anti-MBP), but it seems not to 
bind to the amylose column, it just washes off in the first washing steps. 

We even tried purifying the MBP*2 protein from NEB, diluted in the 
recommended column buffer (20 mM tris / 200 mM NaCl / 1 mM EDTA / 10 mM 
mercapto / pH 7.4), but even then no success. 

I tried raising the NaCl conc. to 500 mM, but no improvement was seen.

Any suggestions are very welcome

Best regards
Maikel



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