Molecular characterization of congenital hyperinsulinism/hyperammonemia caused by glutamate dehydrogenase gene defects

先天性高胰岛素血症和高血氨症（CHH）是由谷氨酸脱氢酶（GDH）失调引起的。我们的特点GDH基因在两个日本CHH患者。与患者2相比，患者1表现出迟发性和轻度低血糖发作以及轻度高氨血症。在淋巴母细胞的GDH活性中，患者1表现出比对照受试者高2倍的基础GDH活性和对GTP抑制的轻度不敏感性。患者2对GTP抑制表现出严重的不敏感性，并且在对照组中对ADP表现出类似的变构刺激。遗传学研究分别在患者1和2中鉴定了杂合和新发L413 V和G446 D突变。COS细胞表达研究证实这两个突变均为致病基因。在COS细胞表达系统中，由于突变GDH基因的剂量效应，L413 V和G446 D对GTP抑制不敏感。生化和遗传学结果表明，L413 V的GDH损伤小于G446 D，与临床表型一致。根据牛GDH的结构，G446 D位于枢螺旋及其周围的GTP结合位点，而L413 V位于触角样结构的α-螺旋。这些不同的突变位点对GDH酶有不同的影响。触角样结构在GDH活性中起重要作用。

Congenital hyperinsulinism and hyperammonemia (CHH) is caused by dysregulation of glutamate dehydrogenase (GDH). We characterized the GDH gene in two Japanese patients with CHH. Patient 1 showed late-onset and mild hypoglycemic episodes and mild hyperammonemia, compared with patient 2. In GDH activity of lymphoblasts, patient 1 showed 2-fold higher basal GDH activity than control subjects and mild insensitivity for GTP inhibition. Patient 2 showed severe insensitivity for GTP inhibition, and similar allosteric stimulation by ADP in the controls. Genetic studies identified heterozygous and de novo L413V and G446D mutations in patients 1 and 2, respectively. COS cell expression study confirmed that both mutations were disease-causing gene. The insensitivity for GTP inhibition in L413V and G446D was emphasized in COS cell expression system as a result of the dosage effect of mutant GDH gene. L413V showed less impairment of GDH than G446D based on biochemical and genetic results, which was consistent with the clinical phenotype. Based on the structure of bovine GDH, G446D was located in GTP binding site of pivot helix and its surroundings, while L413V was located in α-helix of antenna-like structure. These different locations of mutations gave different effects on GDH enzyme. The antenna-like structure plays an important role in GDH activity.