Sortase A Assay Kit

Catalog #

79937

$545 *

Size: 96 reactions

Qty

Bulk/Custom

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*US Pricing only. For international pricing, please contact your local distributor.

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Description

The Sortase A Assay Kit is a fluorogenic assay kit designed to measure Sortase A proteolytic activity for screening and profiling applications.

Figure 1: Illustration of the assay principle.

The fluorescence emitted by donor DNP is quenched due to the proximity of the Abz acceptor in the intact peptide. The protease cleaves the peptide between T (threonine) and G (glycine) and forms an intermediate complex between the cysteine residue within the active site and the C-terminus of the peptide substrate. The cleaved substrate is then covalently bounded to the triglycine nucleophile. The glycine-DNP fragment is released, freeing the emission of fluorescence by DNP.

Need us to run inhibitor screens or profile your compounds against Sortase A? Check out our Protease Screening Services.

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Product Data Gallery

Inhibition of Sortase A activity

Product Info

Storage and Usage

Citations

Assay Kit Format

Fluorogenic

Supplied As

The Sortase A Assay Kit comes in a convenient 96-well format, with enough purified recombinant Sortase A, Triglycine, Abz/DNP substrate, and assay buffer for 96 enzyme reactions.

Materials Required But Not Supplied

Microplate reader capable of reading fluorescence
Adjustable micropipettor and sterile tips
30°C incubator

Format

Catalog #	Name	Amount	Storage
71086	Sortase A, His-Tag*	12.5 μg	-80°C	Avoid multiple freeze/ thaw cycles!
79938	2x Sortase assay buffer	2.5 ml	-20°C
79939	Triglycine	250 μl	-20°C
79940	Abz/Dnp substrate	250 μl	-20°C
79685	Low binding, black NUNC 96-well plate	1	Room Temp

*The concentration of Sortase A is lot-specific and will be indicated on the tube containing the enzyme.

UniProt #

Q9S446

Background

Staphylococcal Sortase A is a bacterial transpeptidase that covalently attaches proteins to the bacterial cell wall, maintaining bacterial virulence and infectivity. Sortase A cleaves a specific peptide sequence (LPXTG recognition motif) within a target protein between threonine and glycine. The cysteine residue of the active site forms a transient thioacyl intermediate complex with the substrate protein. This intermediate complex and substrate is then immediately attacked by oligo-glycine nucleophiles present on peptide-glycans of the bacterial wall to form an amide bond.

Since Sortase A is a critical enzyme needed to maintain the infectivity of the bacterium, it represents a promising therapeutic target. Especially for the treatment of infections caused by antibiotic-resistant bacteria.

References

1. Mazmanian, S.K., et al. 1999. "Staphylococcus aureus sortase, an enzyme that anchors surface proteins to the cell wall." Science 285(5428): 760-763.
2. Spirig, T., et al.. 2011. "Sortase enzymes in Gram‐positive bacteria." Molecular microbiology 82(5): 1044-1059.

Storage/Stability

This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.

Applications

Screen small molecules inhibitors of Sortase A in high-throughput applications
Titer inhibitors of Sortase A

Shipping Temperature

-80°C

Warnings

Avoid freeze/thaw cycles

Disclaimers and Limitations

BPS Bioscience assay kits are validated using the listed components according to our specific protocol. Any deviations to kit components or protocols, such as using different proteins, cell/tissue lysates, or buffers (including using the same component from other commercial sources) will void any warranties on the performance of the assay kit and is not recommended.

End-users should immediately report any issues to [email protected] upon receipt of the kit, such as missing components, damaged vials, no dry ice, etc.

Scientific Category

Sortase

Regulatory Status

Research Use Only