GLUTARYL-COA DEHYDROGENASE AND NEUROLOGIC DISEASE

戊二酰辅酶A脱氢酶与神经系统疾病

• 批准号: 6477147
• 负责人: FRANK E FRERMAN
• 金额: $ 26.99万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-12-10 至 2003-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6477147
• 关键词: X ray crystallography; acidosis; binding sites; brain metabolism; carboxylation; catalyst; chemical binding; chemical kinetics; chemical stability; computer data analysis; enzyme complex; enzyme mechanism; enzyme structure; flavoproteins; gene expression; genetic disorder; human genetic material tag; human tissue; inborn metabolism disorder; molecular assembly /self assembly; neuropathology; oxidation; oxidoreductase; point mutation; protonation

Defects in the flavoprotein, glutaryl-CoA dehydrogenase (GCD), cause glutaric aciduria type I (GA1), an autosomal recessively inherited neurometabolic disorder. GCD catalyzes the alpha, beta dehydrogenation of glutaryl-CoA by a mechanism which is common along acyl-CoA dehydrogenases. GCD also catalyzes the decarboxylation of the enzyme- bound intermediate, glutaconyl-CoA, to crotonyl-CoA and CO2. Decarboxylation of glutaconyl-CoA requires oxidation of the dehydrogenase flavin and protonation of the proposed crotonyl-CoA anion (-CH2=-CH=-CH=-COSCoA). In patients with defects in GCD, the onset of neurological symptoms in GA1 patients usually follows a viral injection early in life. Over 50 missense mutations have been identified that may affect oxidation and decarboxylation of glutaryl-CoA. These mutations may also affect assembly or stability of the tetramer, the oxidation-reduction potential of the dehydrogenase flavin or reoxidation of the dehydrogenase flavin by electron transfer flavoprotein (ETF). The proposed research has the following specific aims. [1] A number of mutant alleles will be expressed and the defective proteins characterized by kinetic and redox methods to access the basis of the enzymatic defects. [2] We will investigate the coupling of glutaryl-CoA oxidation with decarboxylation/protonation of the enzyme bound intermediate, glutaconyl-CoA, using site directed mutations hypothesized to uncouple these steps in catalysis. The crystal structure of GCD with a bound glutaryl-CoA analog and with the reaction intermediate, glutaconyl-CoA, will be determined to provide insight into the decarboxylation reaction. [3] We will investigate the decarboxylation of glutaconyl-CoA directly and define the roles of specific amino acids in the reaction. The participation of the 2'-hydroxyl of the ribityl side chain of the FAD prosthetic group in stabilization of decarboxylation/protonation intermediates will also be determined.

黄蛋蛋白，谷胱甘肽-COA脱氢酶（GCD）的缺陷导致谷氨酸I型I型（GA1），一种常染色体遗传遗传遗传的神经代谢障碍。 GCD通过沿酰基-COA脱氢酶常见的机制催化α-COA的α，β脱氢。 GCD还催化酶结合的中间体的脱羧为谷胱甘肽-COA，催化为crotonyl-COA和CO2。谷氨酸-COA的脱羧需要氧化脱氢酶黄素和所提出的crotonyl-COA阴离子的质子化（-CH2 = -CH = -CH = -CH = -COSCOA）。在GCD缺陷的患者中，GA1患者的神经系统症状的发作通常在生命的早期病毒注射后。已经确定了50多个错义突变，可能会影响谷胱甘肽-COA的氧化和脱羧。这些突变还可能影响四聚体的组装或稳定性，脱氢酶黄素的氧化还原潜力或通过电子转移黄酮（ETF）对脱氢酶黄素的再氧化。拟议的研究具有以下特定目标。 [1]将表达许多突变等位基因，并以动力学和氧化还原方法为特征的有缺陷的蛋白质访问酶促缺陷的基础。 [2]我们将使用位点的定向突变，以使这些步骤在催化中取消这些步骤，研究酶结合的中间体谷氨酸-COA的脱羧/质子化的偶联。 GCD的晶体结构具有结合的谷胱甘肽-COA类似物以及中间体谷氨酰胺-COA的晶体结构，将确定以深入了解脱羧反应。 [3]我们将直接研究谷氨酰胺-COA的脱羧化，并定义特定氨基酸在反应中的作用。还将确定FAD假体基团的肋骨侧链的2'-羟基参与脱羧/质子化中间体的稳定。