I understood that these proteases are very efficient in cleavage. One time, I 
had a construct MBP-3C-protein, I never able to cleave this particular 
construct while I could do well with other truncations.The MBP you are seeing 
might be co-purified with DHBx and unless gel band intensity suggest that its 
coming from fusion protein. I feel cleavage site may not be  accessible for the 
protease. Might be adding few more residues might help. 
Good luckSDY

Date: Sun, 4 Nov 2012 07:24:42 -0800
From: rna19792...@yahoo.com
Subject: [ccp4bb] protein cleavage

Dear CCP4
     I am having a problem with cleaving my fusion protein and I would be 
grateful if you advice me regarding this situation,  I have an MBP-DHBx fusion 
protein and I am trying to cleave it using TEV protease, I have tried different 
ratios and different temperatures  with different incubation time but still it 
will not cleave, all I observe on the gel is the bands of the fusion protein 
which is 59kDa and the MBP which is 42kDa and the TEV protease which is 27kDa 
and no sign of the DHBx which is 17kDa,I have also checked the sequence if 
there was any problem but I could not find anything unusual the sequence was 
fine ,
 so if you have any suggestions regarding this situation I will be thankful 
Best Regards

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