Human Alanine-Glyoxylate Aminotransferase 2 Lowers Asymmetric Dimethylarginine and Protects from Inhibition of Nitric Oxide Production
Human Alanine-Glyoxylate Aminotransferase 2 Lowers Asymmetric Dimethylarginine and Protects from Inhibition of Nitric Oxide Production
复制标题
DOI：
10.1074/jbc.m109.091280
复制
发表时间：
2010-02-19
影响因子：
4.8
通讯作者：
Lentz, Steven R.
中科院分区：
文献类型：
--
作者：
Rodionov, Roman N.;Murry, Daryl J.;Lentz, Steven R.
不对称二甲基精氨酸 (ADMA) 是一氧化氮 (NO) 合酶的内源性抑制剂，其血液浓度升高与糖尿病、高血压、充血性心力衰竭和动脉粥样硬化有关。 ADMA 水平由二甲基精氨酸二甲氨基水解酶 (DDAH) 控制，DDAH 是一种胞质酶，可将 ADMA 水解为瓜氨酸和二甲胺。还提出 ADMA 通过丙氨酸乙醛酸转氨酶 2 (AGXT2)（一种主要在肾脏中表达的线粒体转氨酶）的替代途径进行调节。本研究的目的是确定人 AGXT2 的亚细胞定位，并测试人 AGXT2 过度表达可防止 ADMA 诱导的一氧化氮 (NO) 产生抑制的假设。 AGXT2 从人肾 cDNA 中克隆，并在 COS-7 细胞和人脐静脉内皮细胞中过表达，并带有 C 端 FLAG 表位标签。通过共聚焦显微镜证明了人 AGXT2 的线粒体定位，并通过成熟蛋白的 N 端测序描绘了 41 个氨基酸的 N 端线粒体切割序列。使用腺病毒表达载体在 C57BL/6 小鼠肝脏中过度表达人 AGXT2，导致血浆和肝脏中 ADMA 水平显着降低。人 AGXT2 的过度表达还可以保护内皮细胞免受 ADMA 介导的 NO 产生抑制。我们得出结论，线粒体定位的人 AGXT2 能够在体内有效代谢 ADMA，从而降低 ADMA 水平并改善内皮 NO 产生。
Elevated blood concentrations of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric-oxide (NO) synthase, are found in association with diabetes, hypertension, congestive heart failure, and atherosclerosis. ADMA levels are controlled by dimethylarginine dimethylaminohydrolases (DDAHs), cytosolic enzymes that hydrolyze ADMA to citrulline and dimethylamine. ADMA also has been proposed to be regulated through an alternative pathway by alanine-glyoxylate aminotransferase 2 (AGXT2), a mitochondrial aminotransferase expressed primarily in the kidney. The goal of this study was to define the subcellular localization of human AGXT2 and test the hypothesis that overexpression of human AGXT2 protects from ADMA-induced inhibition in nitric oxide (NO) production. AGXT2 was cloned from human kidney cDNA and overexpressed in COS-7 cells and human umbilical vein endothelial cells with a C-terminal FLAG epitope tag. Mitochondrial localization of human AGXT2 was demonstrated by confocal microscopy and a 41-amino acid N-terminal mitochondrial cleavage sequence was delineated by N-terminal sequencing of the mature protein. Overexpression of human AGXT2 in the liver of C57BL/6 mice using an adenoviral expression vector produced significant decreases in ADMA levels in plasma and liver. Overexpression of human AGXT2 also protected endothelial cells from ADMA-mediated inhibition of NO production. We conclude that mitochondrially localized human AGXT2 is able to effectively metabolize ADMA in vivo resulting in decreased ADMA levels and improved endothelial NO production.