Fluorogenic SIRT substrate 1

Catalog #: 50080
Size: 500 nmole
$230
Description:
A fluorogenic, acetylated peptide substrate for SIRTs. It is a suitable substrate for SIRT1, SIRT2 and SIRT3 (tested so far). The substrate is based on residues 379-382 (Arg-His-Lys-Lys) of p53, a site of regulatory acetylation by the p300 and CBP acetyltransferases on lysines 381 and 382. This substrate fluoresces at excitation 350-380 nm, emission at 440-460 nm. The substrate should be used in conjunction with SIRT Developer.
Supplied As:
5 mM DMSO solution (100 µl)
Format:
DMSO solution
Instructions for use:
Materials :
SIRT Assay buffer (BPS catalog #50090)
SIRT Assay developer (BPS catalog #50089)
SIRT Substrate (BPS catalog #50080)
SIRT1 (1 µg/µl) (BPS catalog #50012) or other SIRT enzyme

Step 1 :
Add the following reagents to a low binding NUNC black plate (VWR catalog #62408-936):
SIRT Assay Buffer, 35 µl
1 mg/ml BSA, 5 µl
200 µM SIRT substrate, 5 µl
SIRT1, 5 µl
Incubate at 37°C for 30 min.

Step 2 :
Stop the reaction by adding 50 µl of SIRT assay developer (undiluted) and incubate the plate at room temperature for 15 min.

Step 3 :
Read samples in a microtiter-plate reading fluorimeter capable of excitation at 350-380 nm and emission at 440-460 nm. “Blank” value (no enzyme negative control) is subtracted from all other values.
Storage / Stability:

Stable at least 12 months from date of receipt, when stored as directed

Application(s):
Study enzyme kinetics, and screen small molecule sirtuin inhibitors for drug discovery and HTS applications.
Reference(s):

1. A. Ito et al. (2001) EMBO J. 20 1331.
2. A. Ito et al. (2002) EMBO J. 21 6236.
3. N.A. Barlev et al. (2001) Mol. Cell 8 1243.

Application Reference:
SIRT2 Interacts with β-Catenin to Inhibit Wnt Signaling Output in Response to Radiation-Induced Stress (2014)

Warning(s):
Protect from direct exposure to light. Avoid freeze/thaw cycles.
Scientific Category:
Deacetylase
Product Type:
Substrate
Data shown is lot-specific. Contact us for specific information on other lots.

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