Molecular analysis of NPC1 and NPC2 gene in 34 Niemann-Pick C Italian Patients: identification and structural modeling of novel mutations
Molecular analysis of NPC1 and NPC2 gene in 34 Niemann-Pick C Italian Patients: identification and structural modeling of novel mutations
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DOI：
10.1007/s10048-009-0175-3
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发表时间：
2009-07-01
期刊：
影响因子：
2.2
通讯作者：
Filocamo, Mirella
中科院分区：
文献类型：
--
作者：
Fancello, Tatiana;Dardis, Andrea;Filocamo, Mirella
Niemann-Pick C 是一种常染色体隐性遗传神经内脏疾病，由内体-溶酶体途径内胆固醇运输失败引起，是由于 NPC1 或 NPC2 基因突变所致。我们对 34 名无关患者进行了特征分析，其中包括 32 名 NPC1 基因突变患者和 2 名 NPC2 基因突变患者。总体而言，遇到了 33 种不同的基因型。在21个未发表的NPC1等位基因中，15个是由于点突变导致13个密码子替换（p.C100S、p.P237L、p.R389L、p.L472H、p.Y634C、p.S636F、p.V780G、p.Q921P、p.Y1019C、p.R1077Q、 p.L1102F、p.A1187V 和 p.L1191F) 和两个提前终止密码子 (p.R934X 和 p.Q447X)；一种新突变体携带两个顺式突变 p.[L648H;M1142T] 和其他四个 NPC1 等位基因，它们是小缺失/插入，导致移码和过早蛋白质截断（p.C31WfsX26、p.F284LfsX26、p.E1188fsX54 和 p.T1205NfsX53）。最后，内含子 4 3' 受体剪接位点的新内含子 c.464-2A > C 变化影响了 NPC1 信使 RNA 加工。我们还发现了由第一个密码子（p.M1L）的变化引起的新的NPC2突变体。通过两种生物信息学方法进一步研究了新的错义突变。豹蛋白分类系统通过计算预测了进化保守位置发生的所有新错义突变的有害影响。另一种生物信息学方法基于对 NPC1 原子模型上错义突变引起的结构改变的预测。计算机分析预测了大多数错义突变的蛋白质功能障碍和/或局部折叠改变。此外，通过SSD模型的原子坐标与胆固醇之间的对接模拟，评估了错义突变（p.Y634C、p.S636F、p.L648H和p.V780G）对甾醇感应结构域（SSD）的影响。
Niemann-Pick C, the autosomal recessive neuro-visceral disease resulting from a failure of cholesterol trafficking within the endosomal-lysosomal pathway, is due to mutations in NPC1 or NPC2 genes. We characterized 34 unrelated patients including 32 patients with mutations in NPC1 gene and two patients in NPC2 gene. Overall, 33 distinct genotypes were encountered. Among the 21 unpublished NPC1 alleles, 15 were due to point mutations resulting in 13 codon replacements (p.C100S, p.P237L, p.R389L, p.L472H, p.Y634C, p.S636F, p.V780G, p.Q921P, p.Y1019C, p.R1077Q, p.L1102F, p.A1187V, and p.L1191F) and in two premature stop codons (p.R934X and p.Q447X); a new mutant carried two in cis mutations, p.[L648H;M1142T] and four other NPC1 alleles were small deletions/insertions leading both to frame shifts and premature protein truncations (p.C31WfsX26, p.F284LfsX26, p.E1188fsX54, and p.T1205NfsX53). Finally, the new intronic c.464-2A > C change at the 3' acceptor splice site of intron 4 affected NPC1 messenger RNA processing. We also found a new NPC2 mutant caused by a change of the first codon (p.M1L). The novel missense mutations were further investigated by two bioinformatics approaches. Panther proein classification system computationally predicted the detrimental effect of all new missense mutations occurring at evolutionary conserved positions. The other bioinformatics approach was based on prediction of structural alterations induced by missense mutations on the NPC1 atomic models. The in silico analysis predicted protein malfunctioning and/or local folding alteration for most missense mutations. Moreover, the effects of the missense mutations (p.Y634C, p.S636F, p.L648H, and p.V780G) affecting the sterol-sensing domain (SSD) were evaluated by docking simulation between the atomic coordinates of SSD model and cholesterol.