第三B型黏多醣儲積症(MPS ⅢB)

Abstract

本研究的目的在分析臺灣四個MPS ⅢB家族的分子致因。聚合酶鏈反應(PCR)放大包含患者NAGLU基因各表現子(exon)片段，經單股核酸構形多型性(SSCP)分析及異偶合(heteroduplex)分析，篩選可能帶有突變的表現子，進行DNA定序，以檢視患者的基因突變。結果發現患者773為複異型合子的突變，其表現子上第220個胺基酸密碼的第三個位置缺失核苷酸C (660delC突變)，表現子6上第565個胺基酸密碼發生CGG→TGG的改變，即由精胺酸Arg轉變成色胺酸Trp(R565W突變)。對偶基因專一的寡核苷酸引子(ASO)雜合的檢測試驗顯示，患者的660delC突變係遺傳白父親，R565W突變則遺傳自母親。患者1146為同型合子的突變，其表現子6上第的6個胺基酸密碼發生CGA→TGA的改變，即由精胺酸轉變成終止密碼(R626X突變)。此變異產生一新的限制蘑DdeI切點，DdeI切割的檢測試驗顯示患者的父親為R626X突變的異型合子。患者1362、1363兄弟皆為同型合子的突變，其表現子2上第130個胺基酸密碼發生CGC→TGC的改變，即由精胺酸轉變成半胱胺酸Cys(R130C突變)。此核苷酸變異使限制酶KpnI的切割點消失，KpnI切割的檢測試驗顯示患者的母親為Rl30C突變的異型合子。患者1377亦為同型合子的突變，其表現子2上第154個胺基酸密碼發生ATA→AGA的改變，即由異白胺酸Il巳轉變成精胺酸(Il54R突變)。此變異經誤配(mismatch)引子的PCR可產生一新的限制臨酶HinfI切點，HinfI切割的檢測試驗顯示，患者的父母親皆為Il54R突變的異型合子。

Molecular lesions of four Chinese families with mucopolysaccharidosis type ⅢB (MPS ⅢB) were investigated. The coding sequences of the NAGLU gene were amplified and examined by single strand conformation polymorphism, heteroduplex, and DNA sequence analyses. Patient 773 has heterozygous mutations; one NAGLU allele has R565W (C-T transition in codon 565) and the other NAGLU allele has 660deiC (deletion of nucleotide Cat cDNA 660). By allele specific oligonucleotide hybridization analysis, mutation R565W was matemally inherited whereas mutation 660deiC patemally inherited. Patient 1146 is homozygous for mutation R626X (C-T transition in codon 626) . By Ddel restriction analysis, the mutation also appeared in 1146's father. Patients 1362 and 1363 are homozygous for mutation Rl30C ( C-T transition in codon 130) . By KpnI restriction analysis, the mutation also appeared in 1362 and 1363's mother. Patient 1377 is homozygous for mutation I154R (T-G transversion in codon 154) . By PCR with mismatch primer and HinfI restriction analysis, the mutation was inherited from both parents.