Gene delivery corrects N-acetylglutamate synthase deficiency and enables insights in the physiological impact of L-arginine activation of N-acetylglutamate synthase.
Gene delivery corrects N-acetylglutamate synthase deficiency and enables insights in the physiological impact of L-arginine activation of N-acetylglutamate synthase.
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DOI：
10.1038/s41598-021-82994-8
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发表时间：
2021-02-11
影响因子：
4.6
通讯作者：
Caldovic L
中科院分区：
文献类型：
--
作者：
Sonaimuthu P;Senkevitch E;Haskins N;Uapinyoying P;McNutt M;Morizono H;Tuchman M;Caldovic L
尿素循环通过将氨转化为尿素来保护中枢神经系统免受氨毒性。N-乙酰谷氨酸合酶（NAGS）催化N-乙酰谷氨酸的形成，N-乙酰谷氨酸是氨甲酰磷酸合成酶1的必需变构激活剂。哺乳动物NAGS的酶活性在L-精氨酸的存在下加倍，但L-精氨酸激活NAGS的生理意义尚不清楚。NAGS基因敲除（Nags−/−）小鼠是一种诱导性高氨血症动物模型，在不补充N-氨甲酰谷氨酸和L-瓜氨酸（NCG + Cit）的情况下会发生高氨血症。我们使用基于腺相关病毒（AAV）的基因转移来纠正Nags−/−小鼠中的NAGS缺陷，确定了拯救Nags−/−小鼠高氨血症所需的载体剂量，并测量了被拯救动物肝脏中Nags mRNA和NAGS蛋白的表达水平。该方法用于研究L-精氨酸对体内尿素生成的影响，方法是用编码野生型或E354 A突变型小鼠NAGS（mNAGS）的AAV载体处理Nags−/−小鼠，mNAGS不被L-精氨酸激活。表达E354 A mNAGS的Nags−/−小鼠存活，但血浆氨浓度升高，尽管E354 A和野生型mNAGS蛋白水平相似。在NAGS缺乏症患者中鉴定了人NAGS中的相应突变（NP_694551.1：p.E360D），其消除了L-精氨酸的结合和激活。我们的研究结果表明，NAGS缺乏症可以挽救基因治疗，并建议L-精氨酸结合NAGS酶是必不可少的正常尿素生成。
The urea cycle protects the central nervous system from ammonia toxicity by converting ammonia to urea. N-acetylglutamate synthase (NAGS) catalyzes formation of N-acetylglutamate, an essential allosteric activator of carbamylphosphate synthetase 1. Enzymatic activity of mammalian NAGS doubles in the presence of L-arginine, but the physiological significance of NAGS activation by L-arginine has been unknown. The NAGS knockout (Nags−/−) mouse is an animal model of inducible hyperammonemia, which develops hyperammonemia without N-carbamylglutamate and L-citrulline supplementation (NCG + Cit). We used adeno associated virus (AAV) based gene transfer to correct NAGS deficiency in the Nags−/− mice, established the dose of the vector needed to rescue Nags−/− mice from hyperammonemia and measured expression levels of Nags mRNA and NAGS protein in the livers of rescued animals. This methodology was used to investigate the effect of L-arginine on ureagenesis in vivo by treating Nags−/− mice with AAV vectors encoding either wild-type or E354A mutant mouse NAGS (mNAGS), which is not activated by L-arginine. The Nags−/− mice expressing E354A mNAGS were viable but had elevated plasma ammonia concentration despite similar levels of the E354A and wild-type mNAGS proteins. The corresponding mutation in human NAGS (NP_694551.1:p.E360D) that abolishes binding and activation by L-arginine was identified in a patient with NAGS deficiency. Our results show that NAGS deficiency can be rescued by gene therapy, and suggest that L-arginine binding to the NAGS enzyme is essential for normal ureagenesis.
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影响因子：
16
作者：
Hallows WC;Yu W;Smith BC;Devries MK;Ellinger JJ;Someya S;Shortreed MR;Prolla T;Markley JL;Smith LM;Zhao S;Guan KL;Denu JM
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DOI：
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发表时间：
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影响因子：
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作者：
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影响因子：
3.7
作者：
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通讯作者：
Caldovic L