Molecular Basis of Maple Syrup Urine Disease

枫糖尿病（MSUD）是一种常染色体隐性遗传疾病，由于分支链α -酮酸脱氢酶复合物（BCKDH）的E_1 α、E_1 β或E_2亚基缺乏。我们进行了以下研究以寻求msud1的分子基础。我们分离并测序了BCKDH的E_2亚基cDNA插入片段。它由2649个碱基对和1431个开放阅读框组成，可翻译成477个氨基酸，3'-非翻译区有1205个碱基对。推导出的氨基酸残基为含脂酰基结构域、e_3结合结构域和非核心结构域。我们分析了人BCKDH e_1 β亚基的cDNA和基因组cDNA结构。分离的cDNA克隆包含4个核苷酸的5‘未翻译序列，1.176个未翻译序列和169个核苷酸的3’未翻译序列。该cDNA编码一个342个氨基酸的亚基，Mr=37,585，其中包括一个50个氨基酸的先导序列。E_1beta亚基基因长度超过150kb，分为10个外显子。所有的剪接供体和受体位点均符合GT/AG规则。转录起始位点位于起始密码子上游47个碱基处。在帽位上游分别有37个碱基和47个碱基存在“CAAT”bax及其反向补体序列，但不存在类似“TATA”的盒状序列。我们发现了三种不同的突变基因；Mennonite MSUD家族中e_1 α亚基基因T-to-A替换，日本MSUD家族中编码线粒体前导肽的11个碱基对重复序列缺失，以及另一个日本MSUD家族中由于剪切异常导致E_2亚基基因78个碱基对缺失。

Maple syrup urine disease (MSUD) is an autosomal recessive inherited disorder due to a deficiency of any subunits, E_1 alpha, E_1 beta or E_2 of branched chain alpha-ketoacid dehydrogenase complex (BCKDH). We performed following study to seek a molecular basis of the MSUD.1. We isolated and sequenced a cDNA insert of E_2 subunit of BCKDH. It consists of 2,649 base pairs with and open reading frame of 1,431 base pairs, which can be translated into 477 amino acids, and 3'-untranslated region of 1,205 base pairs. The deduced amino acid residues were a lipoyl-bearing domain, a E_3-binding domain and innor core domain.2. We analyzed cDNA and genomic cDNA structure of E_1beta subunit of human BCKDH. Isolated cDNA clone contained a 5'-untranslated sequence of 4 nucleotides, the translated sequence of 1.176 uncleotides and a 3'-untranslated sequence of 169 nucleotides. The cDNA encodes for a 342 amino acid subunit with a Mr=37,585, which includes a leader sequence of 50 amino acids. The gene of E_1beta subunit is over 150 kb long and splits into 10 exons. All of the splice donor and acceptor sites confirm to the GT/AG rule. The transcription initiation site was located 47 base upstream of the initiation codon. A "CAAT" bax and its reverse complement sequence were present at 37 bases and 47 bases upstream from the cap sites, but there was no "TATA" box like sequence.3. We found 3 different mutant genes ; A T-to-A substitution in the E_1alpha subunit gene in Mennonite MSUD, a deletion of an 11-base pair repeat sequence, which encodes a mitochondrial ratgetting leader peptide, in a Japanese MSUD family and a 78 base pair deletion in E_2 subunit genedue to splicing abnormalities in another Japanese MSUD family.