REACTION PRODUCTS OF CYTOSOLIC & MITOCHONDRIAL ACONITASES W/ NITRICOXIDE

线粒体(m-ACON)和胞质(c-ACON)乌头酸酶是铁S 催化柠檬酸和异柠檬酸相互转化的酶 通过必需的中间体顺式乌头酸酶。此外， 载脂蛋白c-ACON通过以下途径发挥铁调节蛋白(Irp1)的功能 与保守的茎环结构、铁调节元件结合 (IRES)发现在一些基因的mRNA的非翻译区 蛋白质，包括铁蛋白和转铁蛋白受体。一直以来 以前的研究表明，乌头酸酶可以被氧化剂灭活 通过从[4Fe-4S]团簇中损失一种特定的铁Fe2 形成了EPR可探测的g=2.02[3Fe-4S]形式。 最近一些研究小组的细胞研究表明， NO的产生会使这两种酶失活，而且，在 A-CON的情况导致RNA结合形式IRP1的增加。 研究了不同提纯条件下NO的厌氧反应。 (低于95%)两种乌头酸酶的EPR波谱形式和 活性4Fe是在失去活性后形成的。校长 观察到NO与不同形式的 这两种乌头酸酶都是“g=2.04”的二亚硝基-铁-硫醇络合物。 用连二亚硫酸盐还原的蛋白质产生D9物种。 在c-ACON与NO、瞬时硫基自由基的失活过程中， 观察到g-平行=2.11，g-垂直=2.03。这一速度 M-ACON或c-ACON的失活不受 精氨酸诺酯作为底物来源时底物的存在 不是的。NO与[3Fe-4S]m-ACON反应生成一个物种，g=2.032， 这只在反应的早期阶段观察到，也就是 暂定为亚硝酸铁组氨酸的D9形式 很复杂。两种乌头酸酶的[4Fe-4S]型失活 超氧阴离子或过亚硝酸根都会产生g=2.02[3Fe-4S] 酵素。

Mitochondrial (m-acon) and cytosolic (c-acon) aconitases are Fe-S enzymes that catalyze the interconversion of citrate and isocitrate via the obligatory intermediate cis-aconitase. In addition, the apo-form of c-acon functions as an iron-regulatory protein (IRP1) by binding to conserved stem-loop structures, iron-regulatory elements (IREs) found in the untranslated regions in the mRNA of a number of proteins, including ferritin and transferrin receptor. It has been previously shown that the aconitases can be inactivated by oxidants through the loss of a specific iron, Fe2, from the [4Fe-4S] cluster with the formation of the EPR-detectable, g=2.02 [3Fe-4S] form. Recent cellular studies by a number of groups have indicated that production of NO inactivates both enzymes, and furthermore, in the case of a-con results in an increase in the RNA binding form, IRP1. We have studied the anaerobic reaction of NO with the various purified (less than 95%) forms of both aconitases by EPR spectroscopy and for the active 4Fe forms by following the loss of activity. The principal end product observed upon reaction of NO with the various forms of both aconitases is the "g=2.04," d7 dinitrosyl-iron-thiol complex of the protein which on reduction with dithionite yields the d9 species. During inactivation of c-acon with NO, a transient thiyl radical, g-parallel=2.11 and g-perpendicular=2.03, is observed. The rate of inactivation of either m-acon or c-acon was not retarded by the presence of substrate when spermineNONOate was used as the source of NO. Reaction of NO with [3Fe-4S] m-acon yields a species, g=2.032, that is observed only during the early part of the reaction, which is tentatively assigned to the d9 form of an iron-nitrosyl histidine complex. Inactivation of the [4Fe-4S] forms of both aconitases by either superoxide anion or peroxynitrite produces the g=2.02 [3Fe-4S] enzymes.