臺灣兩位第一型黏多醣儲積症患者的突變篩檢與記述

Humanα-L-iduronidase gene : mutation screening and characterization in two Taiwainese MPS I patients

陳易申(CHEN I SHEN)

指導教授 ： 李桂楨 博士

國立臺灣師範大學/理學院/生物學系/碩士(1999年)

摘要

本論文的目的在探討臺灣兩位第一型黏多醣儲積症 (MPS I) 患者的分子致因。利用聚合酵素鍊反應 (PCR) 放大患者包含 IDUA 基因各表現子 (exon) 及鄰近介入子 (intron) 的片段，進行單股核酸構形多型性 (SSCP) 分析及 DNA 定序分析。結果發現患者 1413 除了在表現子1發生遺傳自母親的 134del12 突變 (秦，1998) 外，尚有一遺傳自父親的 Q584X 突變，即第 1838 個核甘酸發生 C→T 的轉換，造成第 584 個胺基酸由麩醯胺酸 Gln 轉變成終止密碼。患者 174 在表現子 2 上發生 A79V 突變，即第 324 個核甘酸發生 C→T 的轉換，造成第 79 個胺基酸由丙胺酸 Ala 轉變成結頁胺酸 Val；在表現子 14 上則發生 R619G 突變，即第 1943 個核甘酸發生 C →G 的顛換，造成第 619 個胺基酸由精胺酸 Arg 轉變成甘胺酸 Gly。R619G 突變已於先前另一 MPS I 患者 766 的突變分析中被發現且記述 (秦，1998；湯，1998)。進一步構築含 134del12、Q584X、A79V 變異的 IDUA cDNA 重組質體，經 lipofection 的方式轉移至 COS-7 細胞中表現，發現 134del12 重組質體轉移細胞的 IDUA 酵素活性及 mRNA 表現量，和野生型的 IDUA 基因並無明顯差異，但在西方吸漬法卻檢測到大量的 IDUA 前趨蛋白；含 Q584X 變異的 IDUA 酵素活性僅為野生型的 0.4%，mRNA 表現量顯著下降，且未檢測到 IDUA 蛋白質；含 A79V 變異的 IDUA 酵素活性僅為野生型的 0.0%，而 mRNA 表現量雖和野生型無明顯差異，但蛋白質表現量顯著下降。

關鍵字

MPS I

並列摘要

The purpose of this study is to investigate the molecular lesion of two Taiwanese MPS I patients. The coding sequences and exon-intron borders of the IDUA gene were amplified by polymerase chain reaction (PCR) and subjected to single-strand conformation polymorphism (SSCP) and DNA sequencing analyses. The results demonstrated that patient 1413 has heterozygous mutations; the maternal allele has 134del12 (秦，1998) and the paternal allele has Q584X (C-to-T transition in codon 584). Patient 174 has A79V (C-to-T transition in codon 79) in exon 2 and R619G (C-to-G transversion in codon 619) in exon 14. The R619G mutation was identified and characterized previously during mutation sreening in MPS I patient 766 (秦，1998；湯，1998). Expression of 134del12 showed no diffrence in IDUA mRNA level and enzyme activity compared to those of wild type IDUA cDNA upon transfection into COS-7 cells. By western blot analysis, a lot of IDUA precusor protein was detected. In transfected COS-7 cells, Q584X showed traced amounts of α-L-iduronidase activity (0.4% of normal activity). IDUA mRNA level was reduced and no IDUA protein was detected in Q584X transfected COS-7 cells. Expression of A79V showed no appreciable α-L-iduronidase activity (0.0%). Although A79V did not cause apparent reduction in IDUA mRNA level, decreased amount of IDUA protein was observed.

並列關鍵字

第一型黏多醣儲積症

參考文獻

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