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Encompassing Amino Acids: 26-206(end)
Applications: Suitable for use with in vitro protein ligation reactions.
Assay Conditions: Assay was done in the reaction buffer (50 µl) containing 50 mM HEPES (pH=7.4), 150 mM NaCl, 5 mM CaCl2, 5 mM (Gly)3, 25 mM Abz/Dnp substrate, and Sortase A for 30 min at 30°C. Fluorescence intensity was measured at Ex320nm/Em420nm.
Background: Staphylococcal Sortase A is a bacterial transpeptidase that covalently attaches proteins to the bacterial cell wall by cleaving between threonine and glycine at an LPXTG recognition motif and catalyzes the formation of an amide bond between the carboxyl-group of threonine and the amino-group of the cell-wall peptidoglycan. This chemistry has been exploited as a molecular "stapler" to site-specifically link proteins with the C-terminal LPETGX motif to other proteins or molecules possessing a glycine or aminomethylene motif. The Sortase A Pentamutant exhibits higher activity than the wild type protein. This reaction is applicable to proteins in solution and on the living cell surface.
Description: Staphylococcus aureus sortase A Pentamutant (P94R, D160N, D165A, K190E, K196T), a.a. 26-206(end) GenBank Accession No. AF162687 with N-terminal His tag, MW = 21.7 kDa, expressed in an E. coli expression system.
Format: Aqueous buffer solution
Formulation: 40 mM Tris pH 8.0, 110 mM NaCl, 2.2 mM KCl, 400 mM imidazole and 20% glycerol
Genbank: AF162687
Storage Stability: At least 6 months at -80°C.
Tags: N-terminal His-tag
Uniprot: C7TEJ5
Warnings: Avoidfreeze/thaw cycles. Storing diluted enzymeis not recommended, if necessary, usecarrier protein (BSA 0.1 - 0.5%).
Biosafety Level: Not applicable (BSL-1)
References: 1. Chen, I. et al., Proc Natl. Acad. Sci. (USA). 2011; 108(28): 11399-11404.
2. Kappel, K., et al., Protein Sci. 2012 Dec;21(12):1858-71.
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