GLUTARIC ACIDEMIA--MOLECULAR, BIOCHEMICAL, AND CHEMICAL HETEROGENEITY

戊二酸血症(GAI)是人类赖氨酸和色氨酸的先天性错误 氧化导致基底节变性和进行性 儿童时期的运动障碍。死亡通常发生在第一次 十年。这种情况是由于戊二醛-C0A脱氢酶缺乏所致。 (GCDH)，一种含有FAD的线粒体酶，可以氧化 将戊二酰辅酶A脱羧基为巴豆基辅酶A和二氧化碳。我们一直在研究 这种疾病持续了近20年，是第一个描述 并对其器质性酸尿、神经病理学和 酶缺陷。我们克隆并表达了编码人GCDH和 确定了导致GAI的几个突变，目的是学习 这些突变与病理生理学和临床异质性的关系， 以及正常酶和突变酶的催化途径。具体目标为 这一资助期是为了(A)描述导致GAI的GCDH突变，(B) 这些突变与表达的基因的功能异常有关 酶来绘制人GCDH的详细结构-功能图，以及 检测表型和GCDH基因之间的关系以发展 关于导致临床异质性的因素的可检验假设。 在这方面，《特定目的》(C)审查了假设 具有特定剪接位点的GCDH转录本的差异处理 突变是导致临床异质性的一个群体 所有患者都是该突变等位基因的纯合子。须采用的方法 包括GCDH外显子单链构象多态性分析， 突变型GCDH在大肠杆菌中的表达及正常的光谱和动力学研究 和突变型GCDH，以及GAI患者成纤维细胞mRNA的RT-PCR检测。

Glutaric acidemia (GAI) is a human inborn error of lysine and tryptophan oxidation which causes degeneration of the basal ganglia and a progressive movement disorder in childhood. Death usually occurs during the first decade. The condition is due to deficiency of glutaryl-C0A dehydrogenase (GCDH), an FAD- containing mitochondrial enzyme that oxidatively decarboxylates glutaryl-CoA to crotonyl-CoA and CO2. We have been studying this disorder for almost twenty years, being the first to describe the disease and to characterize its organic aciduria, neuropathology, and enzyme defect. We have cloned and expressed cDNA encoding human GCDH and identified several mutations that cause GAI, with the objective of learning how these mutations relate to pathophysiology and clinical heterogeneity, and the catalytic pathways of normal and mutant enzymes. Specific aims for this funding period are to (a) delineate GCDH mutations that cause GAI, (b) correlate these mutations with functional abnormalities in the expressed enzyme to develop a detailed structure-function map of human GCDH, and examine the relations between phenotype and GCDH genotype to develop testable hypotheses about factors contributing to clinical heterogeneity. In this context, specific aim (c) examines the hypothesis t hat differential processing of GCDH transcripts with a specific splice site mutation is responsible for clinical heterogeneity in a population in which all patients are homozygous for this mutant allele. Methods to be employed include single strand conformation polymorphism analysis of GCDH exons, expression of mutant GCDH in E coli, spectral and kinetic studies of normal and mutant GCDH, and RT-PCR of fibroblast mRNA in GAI patients.