Department of Pediatrics, University of British Columbia, Vancouver, BC, Canada.
Mol Genet Metab. 2012 Aug;106(4):478-81. doi: 10.1016/j.ymgme.2012.06.008. Epub 2012 Jun 22.
Pyridoxine dependent epilepsy is an autosomal recessive disorder characterized by early onset seizures responsive to pyridoxine and caused by a defect in the α-aminoadipic semialdehyde dehydrogenase (antiquitin) gene (ALDH7A1). In order to characterize the effects of a series of twelve disease-associated ALDH7A1 missense mutations on antiquitin activity, we generated the mutations in a recombinant human antiquitin cDNA and expressed them in Escherichia coli. We developed an automated spectrophotometric assay of antiquitin enzymatic activity using the natural substrate α-aminoadipic semialdehyde. The substrate was generated using a recombinant lysine aminotransferase gene (lat) from Streptomyces clavuligerus. In the E. coli expression system all the mutants were stably expressed but lacked enzymatic activity. This is consistent with pathogenicity of these mutations in vivo.
吡哆醇依赖型癫痫是一种常染色体隐性遗传病，其特征为早发性癫痫，对吡哆醇有反应，由α-氨基己二酸半醛脱氢酶（旧称 antiquitin）基因（ALDH7A1）缺陷引起。为了研究一系列 12 种与疾病相关的 ALDH7A1 错义突变对 antiquitin 活性的影响，我们在重组人 antiquitin cDNA 中生成这些突变，并在大肠杆菌中表达。我们使用天然底物α-氨基己二酸半醛开发了 antiquitin 酶活性的自动化分光光度测定法。该底物使用来自链霉菌属 clavuligerus 的重组赖氨酸氨基转移酶基因（lat）生成。在大肠杆菌表达系统中，所有突变体均稳定表达，但缺乏酶活性。这与这些突变在体内的致病性一致。
