Molecular identification of aspartate N-acetyltransferase and its mutation in hypoacetylaspartia
Molecular identification of aspartate N-acetyltransferase and its mutation in hypoacetylaspartia
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DOI：
10.1042/bj20091024
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发表时间：
2010-01-01
影响因子：
4.1
通讯作者：
Van Schaftingen, Emile
中科院分区：
文献类型：
--
作者：
Wiame, Elsa;Tyteca, Donatienne;Van Schaftingen, Emile
脑特异性化合物NAA（N-乙酰天冬氨酸）几乎只发生在神经元中，其浓度达到约。通过MRS（磁共振波谱）确定患者中其丰度以评估神经元密度和健康。催化NAA合成的NAT（N-乙酰转移酶）的分子身份仍然未知，因为该酶是膜结合的，难以纯化。数据库检索表明，在人类和小鼠基因组编码的推定NAT（即与已知NAT同源，但具有未表征的催化活性的蛋白质）中，有两种几乎仅在脑中表达，NAT 8L和NAT 14。在HEK-293 T [表达SV 40（猿猴病毒40）的大T抗原的人胚肾-293细胞]中的转染研究表明，NAT 8L催化从L-天冬氨酸和乙酰辅酶A合成NAA，但不催化NAT 14。NAT 8L的特异性、它对天冬氨酸的Km和它对去污剂的敏感性与脑Asp-NAT所描述的那些相似。卵巢）细胞和神经元表达重组NAT 8L表明其与ER（内质网）相关，但不与线粒体相关。在唯一已知缺乏NAA的患者的NAT 8L基因中进行突变搜索，发现存在纯合的19 bp缺失，导致阅读框的改变和功能蛋白的产生缺失。我们的结论是，NAT 8L，神经元特异性蛋白，是负责NAA的合成和突变的主要NAA缺乏症（hypoacetylaspartia）。这种酶的分子鉴定将为阐明这种最丰富的氨基酸衍生物在神经元中的功能和诊断其他患者的低乙酰天冬氨酸提供新的视角。
The brain-specific compound NAA (N-acetylaspartate) occurs almost exclusively in neurons, where its concentration reaches approx. 20 mM. Its abundance is determined in patients by MRS (magnetic resonance spectroscopy) to assess neuronal density and health. The molecular identity of the NAT (N-acetyltransferase) that catalyses NAA synthesis has remained Unknown, because the enzyme is membrane-bound and difficult to purify. Database searches indicated that among putative NATs (i.e. proteins homologous with known NATs, but with uncharacterized catalytic activity) encoded by the human and mouse genomes two were almost exclusively expressed in brain, NAT8L and NAT14. Transfection studies in HEK-293T [human embryonic kidney-293 cells expressing the large T-antigen of SV40 (simian virus 40)] indicated that NAT8L, but riot NAT14, catalysed the synthesis of NAA from L-aspartate and acetyl-CoA. The specificity of NAT8L, its K-m for aspartate and its sensitivity to detergents are similar to those described for brain Asp-NAT. Confocal microscopy analysis of CHO (Chinese-hamster. ovary) cells and neurons expressing recombinant NAT8L indicates that it is associated with the ER (endoplasmic reticulum), but not with mitochondria. A mutation search in the NAT8L gene of the only patient known to be deficient in NAA disclosed the presence of a homozygous 19 bp deletion, resulting in a change in reading frame and the absence of production of a functional protein. We conclude that NAT8L, a neuron-specific protein, is responsible for NAA synthesis and is mutated in primary NAA deficiency (hypoacetylaspartia). The molecular identification of this enzyme will lead to new perspectives in the clarification of the function of this most abundant amino acid derivative in neurons and for the diagnosis of hypoacetylaspartia in other patients.