[Q] Best Protease for Tag Removal
mnoda at lbl.gov
Thu Nov 16 18:35:14 EST 2000
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.
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