Genetic Risk Factors for Hyperhomocysteinemia

高同型半胱氨酸血症的遗传危险因素

• 批准号: 6438897
• 负责人: Roy A GRAVEL
• 金额: $ 28.37万
• 依托单位: UNIVERSITY OF CALGARY
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2006-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6438897
• 关键词: Escherichia coli; aminoacid metabolism; bacterial pigment; betaine compound; biochemical evolution; cardiovascular disorder; cell line; clinical research; coronary disorder; enzyme mechanism; flavodoxin; folate; gene frequency; genetic polymorphism; genetic susceptibility; homocystinuria; human tissue; immunofluorescence technique; in situ hybridization; methionine; methyltransferase; molecular cloning; polymerase chain reaction; western blottings

DESCRIPTION (provided by applicant): It is now well established that elevated total plasma homocysteine is an independent risk factor for cardiovascular disease and that low folate shows association with increased risk for having a child with neural tube defects. Extrapolating from the central role played by a polymorphism of MTHFR in this process, we set out to clone genes and identify polymorphisms to determine if other genes involved in the metabolism of homocysteine would show such associations. A major outcome was the discovery of a novel enzyme methionine synthase reductase (gene and enzyme symbol, MTRR) required for the activation of cobalamin-dependent methionine synthase (MS). Through these experiments we detected a polymorphism, 66A>G (122M), present at a remarkable - aboutO.5 frequency, that appears to be associated with premature coronary artery disease, neural tube defects and Down syndrome. Significant progress included identification of mutations in severely affected patients with MTRR deficiency, modeling the 122M variant in the orthologous E. coli flavodoxin, and development of a mouse model of MTRR deficiency (founder heterozygotes confirmed). We hypothesize that the association of the 122M polymorphism with disease derives from structural and functional disturbances in the variant enzyme with impact on biochemical homeostasis and development. Accordingly, we propose the following specific aims to characterize the mechanism and physiological role of this novel enzyme and polymorphism: (1) Determine the biochemical impact of the 122M mutation in E. coil flavodoxin and humanMTRR. These studies will include biochemical characterization of 122 and M22 MTRR; assessment of FMN binding and domain interactions by 122M flavodoxin; and comparative crystal structure of 122M flavodoxin. (2) Characterize the phenotype of homozygous and heterozygous MTRR knock-out mice to gain understanding of the impact of severe and mild MTRR deficiency on physiology and development. We will examine the expression patterns of MS and MTRR during embryogenesis and in adult tissues by in-situ hybridization, Western blot and tissue immuno-fluorescence; evaluate levels of folate and methionine cycle metabolites in heterozygous and homozygous mutant mice, characterize the general pathology and behavioral phenotype of affected mice; evaluate vascular pathology and function; and explore nutrition-based treatment of homozygous mutant mice. In summary, our study will give a biochemical and physiological face to the impact of 122M on enzyme function, development and disease.

描述（由申请人提供）：现在已经确定了 总血浆同型半胱氨酸是心血管的独立危险因素 疾病和低叶酸表明与患有A的风险增加 神经管缺陷的孩子。从一个扮演的中心角色推断 MTHFR的多态性在此过程中，我们着手克隆基因并识别 多态性以确定其他基因是否参与代谢 同型半胱氨酸将显示出这种关联。一个主要结果是发现 一种新型的蛋氨酸合酶还原酶（基因和酶符号，MTRR） 激活钴胺素依赖性蛋氨酸合酶（MS）所必需的。 通过这些实验，我们检测到存在于66a> g（122m）的多态性 一个显着的 - 大约5频率，似乎与过早有关 冠状动脉疾病，神经管缺陷和唐氏综合症。重要的 进度包括鉴定受影响严重的患者的突变 使用MTRR缺乏，对直系同源大肠杆菌中的122m变体进行建模 黄酮毒素和MTRR缺乏型小鼠模型的发展（创始人 杂合子证实）。我们假设12200万的关联 疾病的多态性来自结构和功能障碍 在变体酶中，对生化稳态和发育产生影响。 因此，我们提出以下特定旨在表征 这种新型酶和多态性的机制和生理作用：（1） 确定122m突变在E. coil黄酮毒素和 HumanMtrr。这些研究将包括122和122的生化特征 M22 MTRR;通过122m黄杜毒素评估FMN结合和结构域相互作用； 和122m黄酮毒素的比较晶体结构。 （2）表征 纯合子和杂合的MTRR敲除小鼠的表型获得 了解严重和轻度MTRR缺乏对生理的影响 和发展。我们将检查MS和MTRR的表达模式 通过原位杂交，蛋白质印迹和成人组织中的胚胎发生和成人组织 组织免疫荧光；评估叶酸和蛋氨酸周期的水平 杂合和纯合突变小鼠中的代谢产物，表征 受影响小鼠的一般病理和行为表型；评估血管 病理和功能；并探索基于营养的纯合处理 突变小鼠。总而言之，我们的研究将提供生化和生理 面对122M对酶功能，发育和疾病的影响。