Tornheim K, Pang H, Costello C E
J Biol Chem. 1986 Aug 5;261(22):10157-62.
The contribution of the purine nucleotide cycle to renal ammoniagenesis was examined in cortical tubule suspensions prepared from acidotic rats and incubated with [alpha-15N]glutamine, [15N]glutamate, or [15N]aspartate. Labeling of ammonia and adenine nucleotides was determined after enzymatic transformations designed to circumvent the technical problem that 15NH3 and H2O have the same nominal mass. Labeling of the adenine nucleotide was undetectable (less than 10%) even after 1 h of incubation. From the measured concentrations of adenine nucleotides and ammonia and the labeling of the ammonia, the flux through the purine nucleotide cycle was calculated to account for less than 1% of the deamination of alpha-amino groups from all three substrates. The glutamate dehydrogenase reaction is therefore the likely pathway for deamination. The rate of 15NH3 production from [alpha-15N]glutamine was two or three times greater than from added [15N]glutamate, indicating a preference for intracellularly generated glutamate. 15NH3 production from added [15N]aspartate was similar to and perhaps slightly greater than that from added [15N]glutamate.
在由酸中毒大鼠制备的皮质肾小管悬液中加入[α-15N]谷氨酰胺、[15N]谷氨酸或[15N]天冬氨酸，对嘌呤核苷酸循环在肾氨生成中的作用进行了研究。在经过酶促转化后测定了氨和腺嘌呤核苷酸的标记情况，该酶促转化旨在规避15NH3和H2O具有相同标称质量这一技术问题。即使孵育1小时后，腺嘌呤核苷酸的标记也无法检测到（小于10%）。根据所测得的腺嘌呤核苷酸和氨的浓度以及氨的标记情况，计算得出通过嘌呤核苷酸循环的通量占所有三种底物α-氨基脱氨基作用的比例不到1%。因此，谷氨酸脱氢酶反应可能是脱氨基的途径。[α-15N]谷氨酰胺产生15NH3的速率比添加的[15N]谷氨酸产生15NH3的速率高两到三倍，表明更倾向于细胞内生成的谷氨酸。添加的[15N]天冬氨酸产生15NH3的情况与添加的[15N]谷氨酸相似，可能略高于后者。
