Alkyl-dihydroxyacetonephosphate synthase - Fate in peroxisome biogenesis disorders and identification of the point mutation underlying a single enzyme deficiency
Alkyl-dihydroxyacetonephosphate synthase - Fate in peroxisome biogenesis disorders and identification of the point mutation underlying a single enzyme deficiency
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DOI：
10.1074/jbc.273.17.10296
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发表时间：
1998-04-24
影响因子：
4.8
通讯作者：
van den Bosch, H
中科院分区：
文献类型：
--
作者：
de Vet, ECJM;Ijlst, L;van den Bosch, H
过氧化物酶体在醚脂质生物合成中发挥着不可或缺的作用，患有多种过氧化物酶体疾病的患者的成纤维细胞和组织中醚磷脂的缺乏就证明了这一点。烷基二羟基丙酮磷酸合酶是一种在醚磷脂生物合成中发挥关键作用的过氧化物酶体酶，在 N 端可裂解前序列中包含过氧化物酶体靶向信号 2 型。使用针对烷基二羟基丙酮磷酸合酶的多克隆抗血清，在受过氧化物酶体疾病影响的患者的成纤维细胞系中检查了该酶的水平。在 Zellweger 综合征和点状根状软骨发育不良患者的成纤维细胞中发现其水平显着降低，表明由于过氧化物酶体输入缺陷，该酶在细胞质中不稳定。在携带过氧化物酶体靶向信号 1 型蛋白的孤立输入缺陷的新生儿肾上腺脑白质营养不良患者中，检测到烷基二羟丙酮磷酸合酶前体形式的水平与成熟形式相当。在对照成纤维细胞中，与过氧化物酶体内定位一致。一位患有烷基二羟丙酮磷酸 (DHAP) 合酶活性孤立缺陷的患者，该蛋白的水平正常。 cDNA 水平的分析揭示了导致该患者酶中 R419H 取代的错义突变。携带这种突变的重组蛋白在大肠杆菌中的表达产生了一种无活性的酶，而相当的对照重组酶是有活性的，这进一步证明这种取代是导致酶和表型无活性的原因。与该结果一致的是，观察到野生型烷基-DHAP合酶活性可以被精氨酸修饰剂苯基乙二醛失活。当底物棕榈酰-DHAP以饱和浓度存在时，该酶可以有效地防止这种失活。编码人烷基二羟丙酮磷酸合酶的基因被定位在染色体2q31上。
Peroxisomes play an indispensible role in ether lipid biosynthesis as evidenced by the deficiency of ether phospholipids in fibroblasts and tissues from patients suffering from a number of peroxisomal disorders. Alkyl-dihydroxyacetonephosphate synthase, a peroxisomal enzyme playing a key role in the biosynthesis of ether phospholipids, contains the peroxisomal targeting signal type 2 in a N-terminal cleavable presequence. Using a polyclonal antiserum raised against alkyl-dihydroxyacetonephosphate synthase, levels of this enzyme were examined in fibroblast cell lines from patients affected by peroxisomal disorders. Strongly reduced levels were found in fibroblasts of Zellweger syndrome and rhizomelic chondrodysplasia punctata patients, indicating that the enzyme is not stable in the cytoplasm as a result of defective import into peroxisomes, In a neonatal adrenoleukodystrophy patient with an isolated import deficiency of proteins carrying the peroxisomal targeting signal type 1, the precursor form of alkyl-dihydroxyacetonephosphate synthase was detected at a level comparable to that of the mature form in control fibroblasts, in line with an intraperoxisomal localization. A patient with an isolated deficiency in alkyl-dihydroxyacetonephosphate (DHAP) synthase activity had normal levels of this protein. Analysis at the cDNA level revealed a missense mutation leading to a R419H substitution in the enzyme of this patient. Expression of a recombinant protein carrying this mutation in Escherichia coli yielded an inactive enzyme, whereas a comparable control recombinant enzyme was active, providing further proof that this substitution is responsible for the inactivity of the enzyme and the phenotype. In line with this result is the observation that wild-type alkyl-DHAP synthase activity can be inactivated by the arginine-modifying agent phenylglyoxal, The enzyme is efficiently protected against this inactivation when the substrate palmitoyl-DHAP is present at a saturating concentration. The gene encoding human alkyl-dihydroxyacetonephosphate synthase was mapped on chromosome 2q31.