[Q] Best Protease for Tag Removal

Michael Oda mnoda at lbl.gov
Thu Nov 16 18:35:14 EST 2000


Hi,

We are working with a His-tagged recombinant protein and would like to 
cleave the tag off in a specific and cheap manner. We have been using 
FactorXa and have found that the cost of cutting 10+ mg of protein to be 
pretty high. I'm sure there must be a better enzyme out there for our 
needs. We have tried Endopeptidase but it cleaves our target protein at 
a secondary site.

Someone suggested an enzyme called "iZyme". It was an unconfirmed 
reference and I have been unable to identify a source for this enzyme. I 
am hoping there is anyone that has heard of this apparently 
recombinant/cheap/specific enzyme. If so, could you please send me 
information on the supplier and its recognition sequence?

We have also been considering placing the amino acids Asp-Pro with their 
acid labile peptide bond between the His tag and the target protein. 
Does anybody have any experience with this particular peptide bond and 
its acid lability? Fortunately our protein is rather durable and folds 
back to a native conformation after 3M Guanidine treatment and appears 
to be somewhat acid stable.

Any suggestions along these lines would be most appreaciated.

                              Thank you,
                              Michael Oda






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