Tang Nelson L S, Hui Joannie, Yong Collin K K, Wong Lawrence T K, Applegarth Derek A, Vallance Hilary D, Law L K, Fung Simon L M, Mak Tony W L, Sung Y M, Cheung K L, Fok T F
Department of Chemical Pathology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, Hong Kong.
Clin Biochem. 2003 Mar;36(2):145-9. doi: 10.1016/s0009-9120(02)00432-0.
Multiple carboxylase deficiency (MCD, MIM:253270) is a common organic aciduria and caused by deficiency of either biotinidase or holocarboxylase synthetase (HLCS; EC 6.3.4.10). Patients commonly present during early infancy with acute metabolic derangements and severe metabolic acidosis. Recently, a late onset form of HLCS deficiency was also described. The different phenotypes (early and late presenting) may be related to a spectrum of mutations in HLCS gene. Applications of mutation analysis in HLCS had been limited previously by the requirement of cDNA from living tissue for study. We described here a genomic approach for molecular diagnosis of HLCS deficiency which we have used to detect mutations in Chinese patients who had the late-onset form of HLCS deficiency. In addition, a fibroblast cell line with MCD from Coriell Cell repositories was also studied.
Three Chinese patients with late onset HLCS deficiency were studied. The genomic sequence of HLCS was retrieved and newly designed primers were used to cover all coding sequences of the gene. PCR products were analyzed by direct sequencing. Population allelic frequencies of mutations detected were determined by genotyping of control samples by restriction fragment length polymorphism.
We found a recurrent mutation, R508W, in the three unrelated Chinese patients. Two were homozygous for this mutation. The other patient was a compound heterozygote of R508W and a novel mutation, D634N. The results suggest that R508W may be an important and relatively prevalent disease-causing mutation in Chinese MCD patients. A fibroblast cell-line from an African patient revealed an additional novel mutation, R565X and a known mutation, V550M.
R508W is a recurrent mutation in Chinese MCD patients which is associated with the late onset phenotype. This new genomic approach for mutation analysis of HLCS gene provides new opportunities in studies of MCD.
多种羧化酶缺乏症（MCD，MIM：253270）是一种常见的有机酸尿症，由生物素酶或全羧化酶合成酶（HLCS；EC 6.3.4.10）缺乏引起。患者通常在婴儿早期出现急性代谢紊乱和严重代谢性酸中毒。最近，还描述了一种迟发型HLCS缺乏症。不同的表型（早发型和迟发型）可能与HLCS基因中的一系列突变有关。以前，由于需要来自活体组织的cDNA进行研究，HLCS突变分析的应用受到限制。我们在此描述了一种用于HLCS缺乏症分子诊断的基因组方法，我们已使用该方法检测患有迟发型HLCS缺乏症的中国患者中的突变。此外，还研究了来自Coriell细胞库的具有MCD的成纤维细胞系。
研究了3例迟发型HLCS缺乏症的中国患者。检索HLCS的基因组序列，并使用新设计的引物覆盖该基因的所有编码序列。通过直接测序分析PCR产物。通过限制性片段长度多态性对对照样品进行基因分型，确定检测到的突变的群体等位基因频率。
我们在3例无亲缘关系的中国患者中发现了一个复发性突变R508W。其中2例为该突变的纯合子。另1例患者是R508W与一个新突变D634N的复合杂合子。结果表明，R508W可能是中国MCD患者中一个重要且相对常见的致病突变。来自一名非洲患者的成纤维细胞系显示了另一个新突变R565X和一个已知突变V550M。
R508W是中国MCD患者中的一个复发性突变，与迟发型表型相关。这种用于HLCS基因突变分析的新基因组方法为MCD研究提供了新的机会。
