PDEnRich™ Cyclic Nucleotide Phosphodiesterase Enrichment Reagent
Cyclic Nucleotide Phosphodiesterase Enrichment Reagent
- Ratio of Total Phosphodiesterase Activity* relative to total protein content increases 4-10X
- PDE protein recoverable, ~70 µg
- Up to 50X concentration of PDE volume, relative to starting volume can be obtained
- 45 minute, scaleable protocol compatible with functional assays, electrophoresis and Mass Spec
Cyclic nucleotide phosphodiesterases (PDEs) hydrolyze the secondary messengers cyclic AMP (cAMP) and cyclic GMP (cGMP) at their 3'-phosphodiester bond, to yield 5'-adenosine monophosphate (5'AMP or AMP) and 5'-guanosine monophosphate (5'GMP or GMP) respectively. These secondary messengers maintain homeostasis and thus play a pivotal role in regulating cellular pathways.
While inhibition of this class of enzymes has had clinical success, a major challenge in designing inhibitors that specifically inhibit PDE subtypes has been the sequence conservation of the catalytic domain among PDE subfamilies. Thus, new methods that can prospect into the structure and the functional properties of conformational variants of PDEs are urgently needed, as the success of PDE inhibitors will depend upon such characterization. The development of a robust enrichment method would be a crucial first step for drug discovery and biomarker classification within the PDE class. However, classical substrate affinity methods have not evolved because of the instability of the cyclic phospho-ester bond. PDEnRich™ is a new reagent kit used for the enrichment and isolation of cyclic AMP (cAMP) and cyclic GMP (cGMP) phosphodiesterases.
PDEnRich™ was discovered upon screening a silica-based, multi-dimensional surface library and optimized for the enrichment of phosphodiesterase activity*. The standard prep protocol starts with 100 µl of tissue/cell extracts, or approximately 0.5 mg total protein, but the process can be scaled up or down to accommodate different sample volumes and protein concentrations. The kit includes all necessary reagents for immediate use.
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