Pyr-Arg-Thr-Lys-Arg-AMC |
Catalog No.GA23423
One-Click Copy Product Info
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Pyr-Arg-Thr-Lys-Arg-AMC also called pERTKR-AMC, is a pentapeptide fluorescent substrate containing AMC (7-amino-4-methylcoumarin).
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 155575-02-3
Sample solution is provided at 25 µL, 10mM.
Pyr-Arg-Thr-Lys-Arg-AMC also called pERTKR-AMC, is a pentapeptide fluorescent substrate containing AMC (7-amino-4-methylcoumarin). Pyr-Arg-Thr-Lys-Arg-AMC can be efficiently cleaved by furin protease, and is therefore often used as a substrate for measuring furinase activity [1].
References:
[1] Neugebauer WA, Parent A, Yuan XW, Day R. C-terminal amidation on aryl hydrazine resin. Adv Exp Med Biol. 2009;611:371-372.
Furin activity assay [1-3]
This plan is based on literature, please modify it to meet your specific needs.
1. Preparation of substrate working solution
(1) Prepare storage solution: Dissolve Pyr-Arg-Thr-Lys-Arg-AMC in DMSO.
Note: Unused storage solution should be aliquoted and stored in the dark at -20°C or -80°C to avoid repeated freezing and thawing.
(2) Prepare working solution: Dilute the storage solution with a reaction buffer (100mM HEPES buffer (pH=7.0) containing 0.2% Triton X-100, 2mM CaCl2, 0.02% sodium azide and 1mg/ml BSA).
Note: Prepare the working solution fresh and store briefly at 4°C or on ice.
2. Preparation of furin enzyme working solution: Dilute furin enzyme with reaction buffer.
3. Prepare a 96-well plate.
Blank wells: Add Pyr-Arg-Thr-Lys-Arg-AMC substrate at a concentration of 10-20μM.
Reaction wells: Add a mixture of Pyr-Arg-Thr-Lys-Arg-AMC substrate and furin enzyme at concentrations of 10-20μM and 10-20nM, respectively.
Inhibitor control wells (optional, for verifying specificity): A mixture of furin enzyme and inhibitor.
4. Pre-incubate the 96-well plate at 37℃ for 30min.
5. Measure the fluorescence intensity at 440-460nm under 350-360nm excitation using a fluorescence spectrophotometer.
Precautions:
1) Fluorescent substrate has quenching problems. Please avoid light as much as possible to slow down fluorescence quenching.
2) For your safety and health, please wear a lab coat and disposable gloves.
References:
[1] Braun E, Hotter D, Koepke L, et al. Guanylate-Binding Proteins 2 and 5 Exert Broad Antiviral Activity by Inhibiting Furin-Mediated Processing of Viral Envelope Proteins. Cell Rep. 2019;27(7):2092-2104.e10.
[2] Shiryaev SA, Farhy C, Pinto A, et al. Characterization of the Zika virus two-component NS2B-NS3 protease and structure-assisted identification of allosteric small-molecule antagonists. Antiviral Res. 2017;143:218-229.
[3] Ma XR, Alugubelli YR, Ma Y, et al. MPI8 is Potent against SARS-CoV-2 by Inhibiting Dually and Selectively the SARS-CoV-2 Main Protease and the Host Cathepsin L. ChemMedChem. 2022;17(1):e202100456.
| Cas No. | 155575-02-3 | SDF | |
| Formula | C37H57N13O9 | M.Wt | 827.94 |
| Solubility | Soluble in DMSO | Storage | Store at -20°C |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 1.2078 mL | 6.0391 mL | 12.0782 mL |
| 5 mM | 241.6 μL | 1.2078 mL | 2.4156 mL |
| 10 mM | 120.8 μL | 603.9 μL | 1.2078 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
g
μL
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >99.00% Appearance: A solid
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 37 reference(s) in Google Scholar.)