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FAM-cGMP PDE V substrate *Green Fluorescence*

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Physical properties
Molecular weight~800
SolventDMSO
Spectral properties
Correction Factor (260 nm)0.32
Correction Factor (280 nm)0.178
Extinction coefficient (cm -1 M -1)83000
Excitation (nm)493
Emission (nm)517
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22
StorageFreeze (< -15 °C); Minimize light exposure
UNSPSC12352200

OverviewpdfSDSpdfProtocol


Molecular weight
~800
Correction Factor (260 nm)
0.32
Correction Factor (280 nm)
0.178
Extinction coefficient (cm -1 M -1)
83000
Excitation (nm)
493
Emission (nm)
517
This green fluorescent cGMP derivative is a specific substrate for phosphodiesterase (PDE) V. It can be used for assaying PDE V activities or screening PDE V inhibitors in combination with anti-cGMP antibody in a FRET readout or FP format. PDE is a group of enzymes that degrade the second messenger molecules: cyclic nucleotides cAMP and cGMP. They regulate the localization, duration, and amplitude of cyclic nucleotide signaling within subcellular domains. PDEs are therefore important regulators of signal transduction mediated by these second messenger molecules. PDE enzymes are often targets for pharmacological inhibition due to their unique tissue distribution, structural and functional properties. Inhibitors of PDE can prolong or enhance the effects of physiological processes mediated by cAMP or cGMP by inhibition of their degradation by PDE. PDE inhibitors have been identified as new potential therapeutics in areas such as pulmonary arterial hypertension, coronary heart disease, dementia, depression and schizophrenia. For example, Sildenafil (Viagra) is an inhibitor of cGMP-specific PDE V, which enhances the vasodilatory effects of cGMP in the corpus cavernosum, and is used to treat erectile dysfunction.

Platform


Fluorescence microplate reader

Excitation490 nm
Emission525 nm
Cutoff515 nm
Recommended plateSolid black

Example protocol


AT A GLANCE

Important      Following protocol only provides a guideline, and should be modified according to your specific needs.

PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

FAM-cGMP PDE V stock solution (1 mM)
Make a 1 mM stock solution by adding 500 µL of DMSO into the vial of 0.5 umol FAM-cGMP PDE V substrate.

PREPARATION OF WORKING SOLUTION

FAM-Cyclic-3’, 5’-GMP PDE V substrate assay solution (2X)
Make 2X FAM-Cyclic-3’, 5’-GMP PDE V substrate assay solution by diluting 1 mM FAM-Cyclic-3’ ,5’-GMP PDE V substrate stock solution into your PDE buffer (such as 10 mM Tris-HCl, pH 7.4, 10 mM Mg Cl2, 1 mM MnCl2) to make a 200 - 400 nM solution.
Note     Make only sufficient quantity needed for the assay.

SAMPLE EXPERIMENTAL PROTOCOL

  1. Mix equal volume of the PDE V standards or samples with 2X FAM-Cyclic-3’ ,5’-GMP PDE V substrate assay solution, and incubate at room temperature for at least 1 hour.
  2. Monitor the fluorescence polarization at Ex/Em = 490/525 nm. 

Spectrum


Open in Advanced Spectrum Viewer
spectrum

Spectral properties

Correction Factor (260 nm)0.32
Correction Factor (280 nm)0.178
Extinction coefficient (cm -1 M -1)83000
Excitation (nm)493
Emission (nm)517

Product Family


NameExcitation (nm)Emission (nm)Extinction coefficient (cm -1 M -1)Correction Factor (260 nm)Correction Factor (280 nm)
TAMRA-cGMP PDE V substrate *Red Fluorescence*552578900000.320.178

Images


References


View all 34 references: Citation Explorer
Oxidative stress employs phosphatidyl inositol 3-kinase and ERK signalling pathways to activate cAMP phosphodiesterase-4D3 (PDE4D3) through multi-site phosphorylation at Ser239 and Ser579
Authors: Hill EV, Sheppard CL, Cheung YF, Gall I, Krause E, Houslay MD.
Journal: Cell Signal (2006): 2056
Leptin interferes with adrenocorticotropin/3',5'-cyclic adenosine monophosphate (cAMP) signaling, possibly through a Janus kinase 2-phosphatidylinositol 3-kinase/Akt-phosphodiesterase 3-cAMP pathway, to down-regulate cholesterol side-chain cleavage cytochr
Authors: Hsu HT, Chang YC, Chiu YN, Liu CL, Chang KJ, Guo IC.
Journal: J Clin Endocrinol Metab (2006): 2761
Helix-1 of the cAMP-specific phosphodiesterase PDE4A1 regulates its phospholipase-D-dependent redistribution in response to release of Ca2+
Authors: Huston E, Gall I, Houslay TM, Houslay MD.
Journal: J Cell Sci (2006): 3799
TcPDE4, a novel membrane-associated cAMP-specific phosphodiesterase from Trypanosoma cruzi
Authors: Alonso GD, Schoijet AC, Torres HN, Flawia MM.
Journal: Mol Biochem Parasitol (2006): 40
Intracellular targeting of phosphodiesterase-4 underpins compartmentalized cAMP signaling
Authors: Lynch MJ, Hill EV, Houslay MD.
Journal: Curr Top Dev Biol (2006): 225
Compartmentalization of cAMP-Dependent Signaling by Phosphodiesterase-4D Is Involved in the Regulation of Vasopressin-Mediated Water Reabsorption in Renal Principal Cells
Authors: Stefan E, Wiesner B, Baillie GS, Mollajew R, Henn V, Lorenz D, Furkert J, Santamaria K, Nedvetsky P, Hundsrucker C, Beyermann M, Krause E, Pohl P, Gall I, Macintyre AN, Bachmann S, Houslay MD, Rosenthal W, Klussmann E.
Journal: J Am Soc Nephrol. (2006)
PDE7A1, a cAMP-specific phosphodiesterase, inhibits cAMP-dependent protein kinase by a direct interaction with C
Authors: Han P, Sonati P, Rubin C, Michaeli T.
Journal: J Biol Chem (2006): 15050
Importance of cAMP-response element-binding protein in regulation of expression of the murine cyclic nucleotide phosphodiesterase 3B (Pde3b) gene in differentiating 3T3-L1 preadipocytes
Authors: Liu H, Tang JR, Choi YH, Napolitano M, Hockman S, Taira M, Degerman E, Manganiello VC.
Journal: J Biol Chem (2006): 21096
DdPDE4, a novel cAMP-specific phosphodiesterase at the surface of dictyostelium cells
Authors: Bader S, Kortholt A, Snippe H, Van Haastert PJ.
Journal: J Biol Chem (2006): 20018
Phosphodiesterase-5 Gln817 is critical for cGMP, vardenafil, or sildenafil affinity: its orientation impacts cGMP but not cAMP affinity
Authors: Zoraghi R, Corbin JD, Francis SH.
Journal: J Biol Chem (2006): 5553