Discovery of halopyridines as quiescent affinity labels: inactivation of dimethylarginine dimethylaminohydrolase.
Discovery of halopyridines as quiescent affinity labels: inactivation of dimethylarginine dimethylaminohydrolase.
复制标题
DOI：
10.1021/ja109207m
复制
发表时间：
2011-02-09
影响因子：
15
通讯作者：
Fast W
中科院分区：
文献类型：
--
作者：
Johnson CM;Linsky TW;Yoon DW;Person MD;Fast W
为了开发可用于生物学应用的二甲基精氨酸二甲氨基水解酶（DDAH）的新型共价修饰剂，测试了使用高通量筛选鉴定的一组“片段”大小的抑制剂的时间依赖性抑制。一种结构类别的灭活剂，4-卤代吡啶，显示DDAH的时间和浓度依赖性灭活，并且详细表征了一个实例，4-溴-2-甲基吡啶（1）的灭活机制。中性形式的卤代吡啶与过量的谷胱甘肽反应性不强。然而，1容易反应，损失其卤化物，在一个选择性的，共价的和不可逆的方式与DDAH的活性位点Cys 249。该活性位点Cys不是特别反应性的（pKa约为1.5）。8.8)1不含木瓜蛋白酶（Cys pKa ca.≤ 4），表明与许多试剂不同，Cys亲核性不是选择性的主导因素。相反，需要通过第二部分Asp 66来结合和稳定灭活剂的更具反应性的吡啶鎓形式以便于反应。这种限制赋予这些灭活剂独特的选择性。据我们所知，卤代吡啶以前没有被报道为蛋白质修饰剂，因此代表了一类新型的静态亲和标记的第一个例子。
In an effort to develop novel covalent modifiers of dimethylarginine dimethylaminohydrolase (DDAH) that are useful for biological applications, a set of “fragment”-sized inhibitors that were identified using a high-throughput screen are tested for time-dependent inhibition. One structural class of inactivators, 4-halopyridines, show time- and concentration-dependent inactivation of DDAH and the inactivation mechanism of one example, 4-bromo-2-methylpyridine (1), is characterized in detail. The neutral form of halopyridines is not very reactive with excess glutathione. However, 1 readily reacts, with loss of its halide, in a selective, covalent and irreversible manner with the active-site Cys249 of DDAH. This active-site Cys is not particularly reactive (pKa ca. 8.8) and 1 does not inactivate papain (Cys pKa ca. ≤ 4), suggesting that, unlike many reagents, Cys nucleophilicity is not a predominating factor in selectivity. Rather, binding and stabilization of the more reactive pyridinium form of the inactivator by a second moiety, Asp66, is required for facile reaction. This constraint imparts a unique selectivity profile to these inactivators. To our knowledge, halopyridines have not previously been reported as protein modifiers, and therefore represent a first-in-class example of a novel type of quiescent affinity label.
登录
查看更多内容
DOI：
10.1039/c0cc02634d
发表时间：
2010-10-14
期刊：
Chemical communications (Cambridge, England)
影响因子：
--
作者：
Knuckley B;Jones JE;Bachovchin DA;Slack J;Causey CP;Brown SJ;Rosen H;Cravatt BF;Thompson PR
通讯作者：
Thompson PR
影响因子：
37.8
作者：
Stühlinger, MC;Tsao, PS;Cooke, JP
通讯作者：
Cooke, JP
影响因子：
2.9
作者：
Stone, EA;Person, MD;Fast, W
通讯作者：
Fast, W
影响因子：
2.9
作者：
Forbes, Scott P.;Druhan, Lawrence J.;Cardounel, Arturo J.
通讯作者：
Cardounel, Arturo J.
影响因子：
15
作者：
Luo, Y;Knuckley, B;Thompson, PR
通讯作者：
Thompson, PR