The efficient and highly specific removal of His tags offered by the TagZyme system makes it the ideal choice for the generation of proteins free from vector-encoded amino acids
Recombinant enzymes included in the TagZyme system allow the efficient and precise exoproteolytic removal of N-terminal His tags from proteins.
In combination with Ni-NTA technology, the TagZyme System provides high-purity proteins free of vector-encoded amino acids for use in applications that demand an absence of non-specific cleavage, the use of recombinant reagents, and a complete removal of all impurities from the target protein.
Recombinant TagZyme enzymes are available in bulk quantities and also in GMP-grade purity to suit any application. FDA Drug Master Files for Ni-NTA matrices are available for support of regulatory issues and matrices are available in bulk quantities to allow easy process scale up.
The TagZyme System uses three enzymes to remove N-terminal His tags with both efficiency and precision.
DAPase Enzyme (dipeptidyl aminopeptidase I) is an exopeptidase that cleaves dipeptides from the N-terminus of proteins, until an amino acid motif that cannot serve as a DAPase substrate (stop point) is reached.
Some proteins contain intrinsic stop points in their amino acid sequence, but if this is not the case, a stop point motif must be introduced by cloning.
The introduction of a glutamine residue directly in front of the first amino acid of the target protein allows the enzymatic generation of a stop point by Qcyclase enzyme (glutamine cyclotransferase), which catalyzes the formation of pyroglutamate from glutamine.
pGAPase enzyme (pyroglutamyl aminopeptidase) removes N-terminal pyroglutamate residues, giving the target protein free of vector-encoded amino acids.
All three enzymes carry C-terminal His tags that allow their removal from reaction mixtures by subtractive Ni-NTA immobilized-metal affinity chromatography (IMAC).
Qiagen offers two new expression vectors, TagZyme pQE-1 and pQE-2, which greatly facilitate the production of His-tag free proteins using the TagZyme System.
Both encode an N-terminal 6xHis tag that possesses a sequence optimized for DAPase digestion and give excellent protein expression rates.
