Sonoda T, Tatibana M
Department of Biochemistry, Chiba University School of Medicine, Japan.
Biochim Biophys Acta. 1990 Feb 26;1033(2):162-8. doi: 10.1016/0304-4165(90)90007-j.
N-Acetyl-L-glutamate has been examined with regard to its ability to activate carbamoyl phosphate synthetase I (EC 6.3.4.16). Substance(s) inhibitory to carbamoyl phosphate synthetase, present even in the partially purified preparation of rat liver extracts, interfered with the measurement of acetylglutamate. In the experiments using chelating agents, metals were apparently involved in this inhibition. When the partially purified preparation of liver extract was placed on a Chelex 100 column, the inhibitor was eliminated and accurate measurements of acetylglutamate content could be made. Evidence supporting the validity of this improved method is given. A significant difference was observed between acetylglutamate levels determined by the present method and by the one using aminoacylase I (N-acylamino acid amidohydrolase, EC 3.5.1.14) to hydrolyze acetylglutamate followed by assay of the glutamate generated. We searched for the presence of glutamate derivatives other than acetylglutamate. When impure tissue preparations containing acetylglutamate were treated with a commercial preparation of aminoacylase, there was an excess amount of glutamate apparently derived from compounds other than acetylglutamate. This can lead to an overestimation of the tissue levels of acetylglutamate.
已对N-乙酰-L-谷氨酸激活氨甲酰磷酸合成酶I（EC 6.3.4.16）的能力进行了研究。即使在大鼠肝脏提取物的部分纯化制剂中存在的对氨甲酰磷酸合成酶有抑制作用的物质，也干扰了乙酰谷氨酸的测定。在使用螯合剂的实验中，金属显然参与了这种抑制作用。当将肝脏提取物的部分纯化制剂置于Chelex 100柱上时，抑制剂被去除，并且可以准确测量乙酰谷氨酸含量。给出了支持这种改进方法有效性的证据。通过本方法测定的乙酰谷氨酸水平与使用氨基酰化酶I（N-酰基氨基酸酰胺水解酶，EC 3.5.1.14）水解乙酰谷氨酸然后测定生成的谷氨酸的方法所测定的水平之间存在显著差异。我们寻找了除乙酰谷氨酸之外的谷氨酸衍生物的存在情况。当用市售的氨基酰化酶制剂处理含有乙酰谷氨酸的不纯组织制剂时，明显存在过量的谷氨酸，这些谷氨酸显然来自除乙酰谷氨酸之外的化合物。这可能导致对组织中乙酰谷氨酸水平的高估。
