Relative role of the glutaminase, glutamate dehydrogenase, and AMP-deaminase pathways in hepatic ureagenesis: studies with 15N.
Relative role of the glutaminase, glutamate dehydrogenase, and AMP-deaminase pathways in hepatic ureagenesis: studies with 15N.
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谷氨酰胺酶、谷氨酸脱氢酶和 AMP 脱氨酶途径在肝尿素生成中的相对作用：15N 的研究。
DOI：
10.1016/0003-9861(92)90008-k
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发表时间：
1992
影响因子：
3.9
通讯作者：
Yudkoff,M
中科院分区：
文献类型：
--
作者：
Nissim,I;Cattano,C;Nissim,I;Yudkoff,M
我们研究了谷氨酰胺酶与谷氨酸脱氢酶 (GLDH) 和嘌呤核苷酸循环 (PNC) 途径在为尿素合成提供氨方面的相对作用。将分离的大鼠肝细胞在 pH 7.4 和 37°C 的 Krebs 缓冲液中孵育，在存在和不存在 1 mmamino-oxyacetate (AOA) 的情况下，补充有 0.1 mm l-鸟氨酸和 1 mm[2-15N]谷氨酰胺、[5-15N]谷氨酰胺、[15N]天冬氨酸或 [15N]谷氨酸作为唯一标记的氮源。在含有15 N标记前体以及浓度与大鼠血浆相似的另外19个未标记氨基酸的培养基中进行一系列单独的培养。利用GC-MS确定前体产物关系以及15 N标记底物朝向15 NH 3 、腺嘌呤核苷酸的6-氨基([6-15NH 2 ]腺嘌呤)、15 N-氨基酸和[ 15 N]脲的通量。与[15N]天冬氨酸一起孵育40分钟后，单标记尿素和双标记尿素的同位素富集分别过量70和20原子％；对于[15N]谷氨酸，单标记尿素和双标记尿素的这些值分别过量约65和约30原子％。在以[ 15 N]天冬氨酸作为唯一底物的实验中，15 NH 3 富集超过了[6-NH 2 ]腺嘌呤，表明[6-15 NH 2 ]腺嘌呤不可能是15 NH 3 的主要前体。添加AOA抑制了[15N]天冬氨酸形成[15N]谷氨酸、15NH3和双标记尿素。然而，AOA对[6-15NH2]腺嘌呤的产生影响不大。在使用[15N]谷氨酸的实验中，AOA抑制[15N]天冬氨酸和双标记尿素的形成，而15NH3的形成增加。在存在生理氨基酸混合物的情况下，[15N]谷氨酸对尿素-N的贡献不到5%。相比之下，无论孵育介质如何，酰胺和谷氨酰胺的氨基氮贡献了总尿素-N 的约 65%。目前的数据表明，当谷氨酸为唯一底物时，通过 GLDH 的通量在为尿素合成提供 NH3 方面比通过 PNC 的通量更重要。然而，在含有大鼠肝脏体内利用的氨基酸混合物的培养基的实验中，与通过谷氨酰胺酶途径产生的氨的量相比，通过GLDH或PNC产生的NH3的比例可以忽略不计。因此，目前的数据表明，通过谷氨酰胺酶从谷氨酰胺的5-N衍生的氨是肝脏尿素生成的主要氮源。
We have studied the relative roles of the glutaminase versus glutamate dehydrogenase (GLDH) and purine nucleotide cycle (PNC) pathways in furnishing ammonia for urea synthesis. Isolated rat hepatocytes were incubated at pH 7.4 and 37 °C in Krebs buffer supplemented with 0.1 mm l-ornithine and 1 mm[2-15N]glutamine, [5-15N]glutamine, [15N]aspartate, or [15N]glutamate as the sole labeled nitrogen source in the presence and absence of 1 mmamino-oxyacetate (AOA). A separate series of incubations was carried out in a medium containing either15N-labeled precursor together with an additional 19 unlabeled amino acids at concentrations similar to those of rat plasma. GC-MS was utilized to determine the precursor product relationship and the flux of15N-labeled substrate toward15NH3, the 6-amino group of adenine nucleotides ([6-15NH2]adenine),15N-amino acids, and [15N]urea. Following 40 min incubation with [15N]aspartate the isotopic enrichment of singly and doubly labeled urea was 70 and 20 atom % excess, respectively; with [15N]glutamate these values were ~65 and ~30 atom % excess for singly and doubly labeled urea, respectively. In experiments with [15N]aspartate as a sole substrate15NH3enrichment exceeded that in [6-NH2] adenine, indicating that [6-15NH2]adenine could not be a major precursor to15NH3. Addition of AOA inhibited the formation of [15N]glutamate,15NH3and doubly labeled urea from [15N]aspartate. However, AOA had little effect on [6-15NH2]adenine production. In experiments with [15N]glutamate, AOA inhibited the formation of [15N]aspartate and doubly labeled urea, whereas15NH3formation was increased. In the presence of a physiologic amino acid mixture, [15N]glutamate contributed less than 5% to urea-N. In contrast, the amide and the amino nitrogen of glutamine contributed ~65% of total urea-N regardless of the incubation medium. The current data indicate that when glutamate is a sole substrate the flux through GLDH is more prominent in furnishing NH3for urea synthesis than the flux through the PNC. However, in experiments with medium containing a mixture of amino acids utilized by the rat liverin vivo, the fraction of NH3derived via GLDH or PNC was negligible compared with the amount of ammonia derived via the glutaminase pathway. Therefore, the current data suggest that ammonia derived from 5-N of glutamine via glutaminase is the major source of nitrogen for hepatic ureagenesis.
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影响因子：
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作者：
J M Mourão;J. McGivan;J. Chappell
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影响因子：
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DOI：
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发表时间：
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期刊：
影响因子：
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DOI：
10.1016/0305-0491(81)90288-1
发表时间：
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期刊：
Comparative Biochemistry and Physiology B
影响因子：
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作者：
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通讯作者：
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影响因子：
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