Design and synthesis of C5 methylated L-arginine analogues as active site probes for nitric oxide synthase
Design and synthesis of C5 methylated L-arginine analogues as active site probes for nitric oxide synthase
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DOI：
10.1021/ja0746159
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发表时间：
2007-10-17
影响因子：
15
通讯作者：
Marletta, Michael A.
中科院分区：
文献类型：
--
作者：
Martin, Nathaniel I.;Woodward, Joshua J.;Marletta, Michael A.
一氧化氮(NO)作为一种生物信号分子的作用已被公认。NO是由一氧化氮合酶(NOSS，EC 1.14.13.39)产生的，一氧化氮合酶是一类能够将L-精氨酸和L-瓜氨酸转化为NO的血红素蛋白。尽管与NOSS相关的知识很多，但与这些酶执行的独特氧化化学有关的机制细节仍有待充分阐明。此外，许多疾病状态都与NO产生过多或过少有关，这使得一氧化氮合酶途径成为治疗发展的一个有吸引力的靶点。由于这些原因，能够提供对NO的产生和/或对NOSS的抑制的机械性见解的分子工具仍然令人感兴趣。我们报道了一些新的L-精氨酸类似物的立体定向合成和测试，这些类似物在氨基酸侧链的第5位具有最小的取代和甲基化(这类类似物以前没有报道)。该合成方法采用了一种改进的光解过程，通过在254 nm处照射适当的双酰基过氧化氢前体，以良好的产率获得了所需的非天然氨基酸。使用诱导型一氧化氮合酶亚型(iNOS(Heme))的血红素结构域结构来评估每种化合物与酶活性部位的结合。这些化合物也被作为诱导型一氧化氮合酶亚型的抑制剂或替代底物进行了研究。所获得的结果为了解酶活性部位的空间和立体化学耐受性提供了新的视角。这些发现也进一步支持了先前提出的NOS催化所必需的保守活性部位水分子的作用。
The role of nitric oxide (NO) as a biological signaling molecule is well established. NO is produced by the nitric oxide synthases (NOSs, EC 1.14.13.39), a class of heme proteins capable of converting L-arginin to NO and L-Citrulline. Despite the large body of knowledge associated with the NOSs, mechanistic details relating to the unique oxidative chemistry performed by these enzymes remain to be fully elucidated. Furthermore, a number of disease states are associated with either the over- or underproduction of NO, making the NOS pathway an attractive target for the development of therapeutics. For these reasons, molecular tools capable of providing mechanistic insights into the production of NO and/or the inhibition of the NOSs remain of interest. We report here the stereospecific synthesis and testing of a number of new L-arginine analogues bearing a minimal substitution, methylation at position 5 of the amino acid side chain (such analogues have not been previously reported). The synthetic approach employed a modified photolysis procedure whereby irradiation of the appropriate diacylperoxide precursors at 254 nm gave access to the required unnatural amino acids in good yields. A heme domain construct of the inducible NOS isoform (iNOS(heme)) was used to assess the binding of each compound to the enzyme active site. The compounds were also investigated as either inhibitors of, or alternate substrates for, the inducible NOS isoform. The results obtained provide new insight into the steric and stereochemical tolerance of the enzyme active site. These findings also further support the role of a conserved active site water molecule previously proposed to be necessary for NOS catalysis.