A new nonhydrolyzable reactive cGMP analogue, (Rp)-guanosine-3′,5′-cylic-S-(4-bromo-2,3-dioxobutyl)monophosphorothioate, which targets the cGMP binding site of human platelet PDE3A

被引：3

作者：

Hung, Su H. ^{[1
]}

Liu, Andy H. ^{[1
]}

Pixley, Robin A. ^{[1
]}

Francis, Penelope ^{[1
]}

Williams, LaTeeka D. ^{[1
]}

Matsko, Christopher M. ^{[1
]}

Barnes, Karine D. ^{[1
]}

Sivendran, Sharmila ^{[2
]}

Colman, Roberta F. ^{[2
]}

Colman, Robert W. ^{[1
]}

机构：

[1] Temple Univ, Sch Med, Sol Sherry Thrombosis Res Ctr, Philadelphia, PA 19140 USA

[2] Univ Delaware, Dept Chem & Biochem, Newark, DE 19716 USA

来源：

BIOORGANIC CHEMISTRY | 2008年 / 36卷 / 1-3期

关键词：

affinity label; Rp-cGMPS-BDB; human platelets; phosphodiesterase; PDE3A;

D O I：

10.1016/j.bioorg.2008.02.006

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

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(1) = 43.4 +/- 7.2 mu M 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 mu M) > Sp-cGMPS (124), Sp-cAMPS (182) > GMP (1517), Rp-cAMPS (3762), AMP (4370 mu M). 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. (C) 2008 Elsevier Inc. All rights reserved.

引用

页码：141 / 147

页数：7