molecular basis of argininosuccinate synthetase deficiency in citrullinemia.

瓜氨酸血症精氨基琥珀酸合成酶缺乏的分子基础。

• 批准号: 02670126
• 负责人: KOBAYASHI Keiko
• 金额: $ 1.34万
• 依托单位: Kagoshima University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1991
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02670126/
• 关键词: Citrullinemia; Argininosuccinate synthetase deficiency; Urea cycle enzyme deficiency; Tissue specific abnormality; Homozygosity mapping; RFLP analysis; RFLP 解析; in vitro発現系; PCR; compound heterozygote; 臓器特異的異常

Citrullinemia is an autosomal recessive disease caused by deficiency of argininosuccinate synthetase(ASS)which functions as a member of urea cycle in the liver.This enzyme defect is found in all tissues or cells of the classic neonatal citrullinemia. Fourteen mutations were identified by sequencing amplified CDNA from 14 neonatal patients. Mutations causing neonatal citrullinemia are extremely heterogeneous, and all non-consanguineous individuals studied to date are compound heterozygotes. The nucleotide coding sequence and deduced amino acid analysis are available for four mammalian species, yeast and three bacterial species. Six of nine missense mutations in humans occur in amino acid positions that are completely conserved in these organisms. To confirm these missense and deletion mutations, we established the in vitro expression system for ASS protein using PCAGGS vector in BMT-10 cells. Five mutant CDNA were tested in this expression system and were confirmed that all mutant had no ASS activity and no or low level of ASS-CRM.We also describe a different type of citrullinemia(adult onset form)which occurs in association with decreased ASS activity in the liver but normal activity in the kidney and other cells. Decreased hepatic ASS with normal kinetics properties and heat stability is accompanied by normal levels of ASSmRNA in the liver, no gross structural abnormality, and translational activity. No mutations were found in the whole sequence of ASSmRNA(except 20 base of 5'-end). We have analyzed about 80 patients with adult citrullinemia to date, and the proportion of patients with consanguinity were approximately 20%. RFLP analysis of 11 patients from consanguineous marriage suggested the possibility that the primary defect is not located on the ASS gene. We are now performing further homozygosity mapping.

瓜氨酸血症（Citrullinemia）是一种常染色体隐性遗传病，由肝脏尿素循环中的一个成员--氨基琥珀酸合成酶（ASS）缺陷引起，这种酶缺陷见于经典的新生儿瓜氨酸血症的所有组织或细胞。通过对14例新生儿患者扩增的cDNA进行测序，确定了14个突变。引起新生儿瓜氨酸血症的突变是极其异质的，迄今为止研究的所有非血缘个体都是复合杂合子。核苷酸编码序列和推导的氨基酸分析可用于四种哺乳动物、酵母和三种细菌。人类的九个错义突变中有六个发生在这些生物体中完全保守的氨基酸位置。为了证实这些错义和缺失突变，我们建立了ASS蛋白的体外表达系统，使用PCAGGS载体在BMT-10细胞中。在该表达系统中检测了5个突变体cDNA，证实所有突变体都没有ASS活性，没有或低水平的ASS-CRM。我们还描述了一种不同类型的瓜氨酸血症（成人发病形式），其发生与肝脏中ASS活性降低有关，但在肾脏和其他细胞中的活性正常。具有正常动力学特性和热稳定性的肝脏ASS降低伴随着肝脏中ASSmRNA的正常水平，没有大体结构异常和翻译活性。ASSmRNA全序列（除5 ′端20个碱基外）未发现突变。迄今为止，我们分析了约80例成人瓜氨酸血症患者，近亲患者的比例约为20%。对11例近亲结婚患者的RFLP分析提示原发性缺陷可能不位于ASS基因上。我们现在正在进行进一步的纯合性作图。

项目成果

Osamu Nukada: "A case of citrullinemia with abnormal messenger RNA for Argininosuccinate synthetase." Acta Paediatr.Jpn.33. 672-677 (1991)

Osamu Nukada：“精氨酸琥珀酸合成酶信使 RNA 异常的瓜氨酸血症病例。”

Keiko Kobayashi: "Heterogeneity of mutations in argininosuccinate synthetase causing human citrullinemia" J. Biol. Chem.265. 11361-11367 (1990)

Keiko Kobayashi：“导致人类瓜氨酸血症的精氨酸琥珀酸合成酶突变的异质性”J. Biol。

Keiko Kobayashi: "Simultaneous detection of mutant gene and transgene in ornithine transcarbamylase (OTC)-deficient, spf-ash mice introduced rat OTC gene" J. Inherit. Metab. Dis.(1992)

Keiko Kobayashi：“在鸟氨酸转氨甲酰酶 (OTC) 缺陷、引入大鼠 OTC 基因的 spf-ash 小鼠中同时检测突变基因和转基因” J. Inherit。

小林 圭子ら: "先天性代謝異常症の遺伝子解析と診断" 医学のあゆみ. 153. 522-527 (1990)

Keiko Kobayashi 等人：“先天性代谢紊乱的基因分析和诊断”，医学史 153. 522-527 (1990)。

神代 龍吉: "肝移値を受けた成人型シトルリン血症の1症例" 肝・胆・膵. 20. 843-849 (1990)

Ryukichi Kamishiro：“具有肝脏转移值的成人型瓜氨酸血症病例”《肝脏、胆汁和胰腺》20. 843-849 (1990)。