Anderson P M, Casey C A
J Biol Chem. 1984 Jan 10;259(1):456-62.
High levels of both glutamine synthetase and a unique L-glutamine- and N-acetyl-L-glutamate-dependent carbamoyl phosphate synthetase are present in the mitochondria in livers of marine urea-retaining elasmobranchs (Casey, C. A., and Anderson, P. M. (1982) J. Biol. Chem. 257, 8449-8453). On the basis of these observations it has been suggested that in these species carbamoyl phosphate and, consequently, one of the nitrogen atoms of citrulline and, ultimately, urea, are derived directly from glutamine rather than from ammonia as occurs in mammalian ureotelic species. The purpose of this study was to obtain evidence for this role of glutamine. Isolated hepatic mitochondria from Squalus acanthias incubated with ammonia plus glutamate, ornithine, bicarbonate, inorganic phosphate, and succinate as an energy source were found to synthesize citrulline at a rate comparable to the rate of urea synthesis observed in vivo. Citrulline synthesis proceeds at maximal rates even when the ammonia concentration is as low as 0.05 mM and is stoichiometric with the amount of ammonia initially present. Synthesis from ammonia does proceed in the absence of glutamate, but a much higher concentration of ammonia (congruent to 4 mM) is required to achieve a half-maximal rate. Glutamine can substitute for ammonia plus glutamate as the nitrogen-donating substrate for citrulline synthesis. Selective inhibition of the glutamine-dependent activity of the carbamoyl phosphate synthetase in the isolated mitochondria completely inhibits the ability of the mitochondria to synthesize citrulline from glutamine or from ammonia plus glutamate, whereas selective inhibition of glutamine synthetase inhibits citrulline synthesis from ammonia plus glutamate, but not from glutamine. These observations provide direct evidence that ammonia assimilation for citrulline synthesis (and, therefore, urea synthesis) in these species involves intermediate formation of glutamine.
在海洋尿素保留型板鳃亚纲动物的肝脏线粒体中，谷氨酰胺合成酶以及一种独特的依赖L-谷氨酰胺和N-乙酰-L-谷氨酸的氨甲酰磷酸合成酶的水平都很高（凯西，C.A.，和安德森，P.M.（1982年）《生物化学杂志》257卷，8449 - 8453页）。基于这些观察结果，有人提出在这些物种中，氨甲酰磷酸以及因此瓜氨酸的一个氮原子乃至最终尿素的一个氮原子，直接来源于谷氨酰胺，而不像在哺乳动物排尿素物种中那样来源于氨。本研究的目的是获取谷氨酰胺这一作用的证据。发现用氨加谷氨酸、鸟氨酸、碳酸氢盐、无机磷酸盐以及琥珀酸作为能量来源培养的尖吻棘鲛分离肝线粒体，其瓜氨酸合成速率与体内观察到的尿素合成速率相当。即使氨浓度低至0.05 mM时，瓜氨酸合成仍以最大速率进行，且与最初存在的氨量呈化学计量关系。在没有谷氨酸的情况下，由氨合成瓜氨酸的过程也能进行，但需要高得多的氨浓度（约4 mM）才能达到最大速率的一半。谷氨酰胺可以替代氨加谷氨酸作为瓜氨酸合成的供氮底物。对分离线粒体中氨甲酰磷酸合成酶的谷氨酰胺依赖性活性进行选择性抑制，会完全抑制线粒体从谷氨酰胺或从氨加谷氨酸合成瓜氨酸的能力，而对谷氨酰胺合成酶进行选择性抑制会抑制从氨加谷氨酸合成瓜氨酸，但不抑制从谷氨酰胺合成瓜氨酸。这些观察结果提供了直接证据，表明在这些物种中，用于瓜氨酸合成（进而尿素合成）的氨同化涉及谷氨酰胺的中间形成。
