REACTION PRODUCTS OF CYTOSOLIC & MITOCHONDRIAL ACONITASES W/ NITRICOXIDE

细胞质的反应产物

• 批准号: 6279855
• 负责人: M C KENNEDY
• 金额: $ 0.25万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-04-01 至 1999-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6279855
• 关键词: bioengineering /biomedical engineering; biological products; biomedical resource; proteins

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.

线粒体（m-acon）和细胞溶质（c-acon）乌头酸酶是Fe-S 催化柠檬酸盐和异柠檬酸盐相互转化的酶 通过强制性中间体顺乌头酸酶。 此外该 c-acon的apo形式作为铁调节蛋白（IRP 1）发挥作用， 结合保守的茎环结构，铁调节元件 （IREs）发现在非翻译区的mRNA的数量， 铁蛋白和转铁蛋白受体。 已经 以前表明乌头酸酶可以被氧化剂灭活， 通过从[4Fe-4S]簇中损失特定的铁Fe 2， 形成EPR可检测的g=2.02的[3Fe-4S]形式。 一些研究小组最近的细胞研究表明， NO的产生使这两种酶失活，此外，在 a-con的情况导致RNA结合形式IRP 1的增加。 我们研究了NO的厌氧反应与各种纯化 (less超过95%）的两种乌头酸酶形式， 随着活性的丧失而形成活性4Fe。 校长 在NO与各种形式的 这两种乌头酸酶都是“g=2.04”的d 7二亚硝酰基-铁-硫醇复合物， 该蛋白质在用连二亚硫酸盐还原时产生D9物质。 在c-acon与NO（一种瞬时硫基自由基）的失活过程中， g-平行=2.11和g-垂直=2.03。 率 无论是m-acon还是c-acon的失活， 当精胺NONOate用作底物来源时， 号 NO与[3Fe-4S] m-acon反应产生一种物种，g=2.032， 这只在反应的早期阶段观察到， 暂时归属于亚硝酰组氨酸铁的d9形式 复杂. 两种乌头酸酶的[4Fe-4S]形式的失活 超氧阴离子或过氧亚硝酸根产生g=2.02 [3Fe-4S] 内切酶