| Literature DB >> 18394675 |
Su H Hung1, Andy H Liu, Robin A Pixley, Penelope Francis, LaTeeka D Williams, Christopher M Matsko, Karine D Barnes, Sharmila Sivendran, Roberta F Colman, Robert W Colman.
Abstract
The amino acids involved in substrate (cAMP) binding to human platelet cGMP-inhibited cAMP phosphodiesterase (PDE3A) are identified. Less is known about the inhibitor (cGMP) binding site. We have now synthesized a nonhydrolyzable reactive cGMP analog, Rp-guanosine-3',5'-cyclic-S-(4-bromo-2, 3-dioxobutyl)monophosphorothioate (Rp-cGMPS-BDB). Rp-cGMPS-BDB irreversibly inactivates PDE3A (K(I)=43.4+/-7.2muM and k(cart)=0.007+/-0.0006 min(-1)). The effectiveness of protectants in decreasing the rate of inactivation by Rp-cGMPS-BDB is: Rp-cGMPS (K(d)=72 microM)>Sp-cGMPS (124), Sp-cAMPS (182)>GMP (1517), Rp-cAMPS (3762), AMP (4370 microM). NAD(+), neither a substrate nor an inhibitor of PDE3A, does not protect. Nonhydrolyzable cGMP analogs exhibit greater affinity than the cAMP analogs. These results indicate that Rp-cGMPS-BDB targets favorably the cGMP binding site consistent with a docking model of PDE3A-Rp-cGMPS-BDB active site. We conclude that Rp-cGMPS-BDB is an effective active site-directed affinity label for PDE3A with potential for other cGMP-dependent enzymes.Entities:
Mesh：
Substances：
Year: 2008 PMID： 18394675 PMCID： PMC2516375 DOI： 10.1016/j.bioorg.2008.02.006
Source DB: PubMed Journal: Bioorg Chem ISSN： 0045-2068 Impact factor: 5.275