Inhibition of N-acetylglutamate synthase by various monocarboxylic and dicarboxylic short-chain coenzyme A esters and the production of alternative glutamate esters
Inhibition of N-acetylglutamate synthase by various monocarboxylic and dicarboxylic short-chain coenzyme A esters and the production of alternative glutamate esters
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DOI：
10.1016/j.bbadis.2013.04.027
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发表时间：
2014-12-01
影响因子：
6.2
通讯作者：
Wanders, R. J. A.
中科院分区：
文献类型：
--
作者：
Dercksen, M.;Ijlst, L.;Wanders, R. J. A.
高氨血症是常见的有机酸中毒。一种可能的机制涉及通过短链酰基辅酶A抑制酶N-乙酰谷氨酸合酶（NAGS），所述短链酰基辅酶A由于细胞中氨基酸和/或脂肪酸的缺陷性催化而积累。本研究的目的是研究各种酰基辅酶A对NAGS活性以及谷氨酸酯形成的影响。采用灵敏的酶测定法和超高效液相色谱-串联质谱（UPLC-MS/MS）产物分析法在体外测定NAGS活性。丙酰辅酶A和丁酰辅酶A被证明是N-乙酰谷氨酸（NAG）形成的最有效的抑制剂。支链氨基酸相关的辅酶A（异戊酰辅酶A、3-甲基巴豆酰辅酶A、异丁酰辅酶A）对NAGS的抑制作用不明显，而二羧酸短链酰基辅酶A（甲基丙二酰辅酶A、琥珀酰辅酶A、戊二酰辅酶A）的抑制作用最小。随后的工作表明，最强大的抑制剂也被证明是N-酰基谷氨酸形成的最佳底物。此外，我们确定了N-异戊酰谷氨酸，N-3-甲基巴豆酰谷氨酸和N-异丁酰谷氨酸（后两种微量），在不同的有机酸中毒患者的尿液中。总的来说，这些发现解释了继发性高氨血症的促成因素之一，其导致有机酸代谢中通过尿素循环的体内通量减少，并导致氨消除不足。(C)2013爱思唯尔有限公司版权所有。
Hyperammonemia is a frequent finding in various organic acidemias. One possible mechanism involves the inhibition of the enzyme N-acetylglutamate synthase (NAGS), by short-chain acyl-CoAs which accumulate due to defective catabolism of amino acids and/or fatty acids in the cell. The aim of this study was to investigate the effect of various acyl-CoAs on the activity of NAGS in conjunction with the formation of glutamate esters. NAGS activity was measured in vitro using a sensitive enzyme assay with ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) product analysis. Propionyl-CoA and butyryl-CoA proved to be the most powerful inhibitors of N-acetylglutamate (NAG) formation. Branched-chain amino acid related CoAs (isovaleryl-CoA, 3-methylcrotonyl-CoA, isobutyryl-CoA) showed less pronounced inhibition of NAGS whereas the dicarboxylic short-chain acyl-CoAs (methylmalonyl-CoA, succinyl-CoA, glutaryl-CoA) had the least inhibitory effect. Subsequent work showed that the most powerful inhibitors also proved to be the best substrates in the formation of N-acylglutamates. Furthermore, we identified N-isovalerylglutamate, N-3-methylcrotonylglutamate and N-isobutyrylglutamate (the latter two in trace amounts), in the urines of patients with different organic acidemias. Collectively, these findings explain one of the contributing factors to secondary hyperammonemia, which lead to the reduced in vivo flux through the urea cycle in organic acidemias and result in the inadequate elimination of ammonia. (C) 2013 Elsevier B.V. All rights reserved.