Recombinant Sortase A5 protein

Contents

A representative Technical Data Sheet (TDS) is provided here. Please refer to the lot-specific TDS you will receive with your order for the lot-specific buffer contents and protein concentration.

Background

Sortase belongs to a class of transpeptidases that utilize an active site cysteine thiol to modify proteins by recognizing and cleaving a carboxy-terminal sorting signal, LPXTG (where X is any amino acid), between the threonine and glycine residues. Sortase A5 is an engineered pentamutant variant of the wild-type sortase from Staphylococcus aureus that is significantly more active than the wild-type sortase. Sortase A5 site-specifically labels antibodies or proteins when the LPXTG recognition sequence is displayed. Easily attach a wide variety of labels such as peptides, DNA, carbohydrates or fluorophores containing a poly-Glycine sequence (Gly)_n (where n = 3 or more Glycine residues). Sortase A5 Pentamutant is covered by US patent number 9,267,127.

Application Notes

Sortase A5 recognizes an antibody or protein genetically engineered to contain the LPXTG motif (where X is any amino acid). Sortase A5 cleaves this sequence between the threonine and glycine residues and the terminal glycine is then replaced with any poly-Glycine (G)n label. Sortase A5 is used in Active Motif’s Sortag-IT™ Labeling Kits to attach HRP, biotin, fluorophores and other labels directly to Active Motif’s AbFlex™ recombinant antibodies (rAb). The activity of both wild type and Sortase A5 pentamutant proteins are Ca²⁺ dependent, therefore, Active Motif’s Sortag-IT labeling buffers and reagents are formulated to contain Ca²⁺ for optimal protein labeling. In addition, our AbFlex recombinant antibodies are provided in HEPES buffer, which is does not bind Ca²⁺ and will not interfere with the Sortase A5 enzymatic activity.

Protein Details

Recombinant Sortase A5 protein (S. aureaus, Uniprot A0A077UNB8-1), containing amino acid substitutions P94R, D160N, D165A, K190E and K196T, was expressed in E. coli and includes a C-terminal 6xHis Tag. The molecular weight of the protein is 17.8 kDa.

Protein Sequence: MQAKPQIPKDKSKVAGYIEIPDADIKEPVYPGPATREQLNRGVSFA EENESLDDQNISIAGHTFIDRPNYQFTNLKAAKKGSMVYFKVGNETRKYKMTSIRNVKPT
AVEVLDEQKGKDKQLTLITCDDYNEETGVWETRKIFVATEVKLEHHHHHH

References

The following applications have been published using this protein. Unless noted above, Active Motif may not have validated this product for use in these applications:

Protein activity assay

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Storage

Store at -80°C to prevent degradation and avoid repeated freeze/thaw cycles. This product is guaranteed for 6 months from date of arrival.