Cyr D M, Egan S G, Brini C M, Tremblay G C
Department of Biochemistry and Biophysics, University of Rhode Island, Kingston 02881.
Biochem Pharmacol. 1991 Jul 15;42(3):645-54. doi: 10.1016/0006-2952(91)90328-3.
Synthesis of glucose from lactate and generation of urea from ammonia were inhibited when sodium benzoate was added to suspensions of rat hepatocytes. Assays with isolated mitochondria suggested pyruvate carboxylase and the N-acetyl-L-glutamate (NAG)-dependent carbamoylphosphate synthetase (CPS-I) as potential sites of inhibition for both pathways, owing to a shared dependency on aspartate efflux from the mitochondria and its subsequent conversion to oxaloacetate in the cytosol. Assays with isolated hepatocytes indicated inhibition to be initiated by accumulation of benzoyl CoA with a resultant depletion of free CoA and acetyl CoA. Measurements of adenine nucleotides showed that benzoate metabolism did not sufficiently alter energy status to account for the observed inhibition. Consistent with these interpretations, acceleration of the conversion of benzoyl CoA to hippurate by the addition of glycine restored the levels of free CoA and acetyl CoA and the rates of gluconeogenesis and ureagenesis. Reduction of the levels of aspartate and glutamate, presumably by interference with the anapleurotic function of pyruvate carboxylase, most likely accounted for inhibition of gluconeogenesis by benzoate. Whether reduced flux through the urea cycle also contributed to inhibition of gluconeogenesis (by diminishing cytosolic conversion of aspartate to oxaloacetate) requires further study. Depression of glutamate and acetyl CoA to levels at or below the Km for NAG synthetase probably accounted for the observed inhibition of ureagenesis. Rates of urea production were observed to vary with changes in the levels of NAG, suggesting NAG-dependent CPS-I to be the primary site of inhibition of ureagenesis by benzoate.
当向大鼠肝细胞悬液中添加苯甲酸钠时，乳酸合成葡萄糖以及氨生成尿素的过程受到抑制。对分离的线粒体进行的测定表明，丙酮酸羧化酶和N - 乙酰 - L - 谷氨酸（NAG）依赖性氨甲酰磷酸合成酶（CPS - I）可能是这两条途径的抑制位点，这是因为它们都依赖线粒体天冬氨酸外流以及随后其在胞质溶胶中转化为草酰乙酸。对分离的肝细胞进行的测定表明，抑制作用是由苯甲酰辅酶A的积累引发的，导致游离辅酶A和乙酰辅酶A的消耗。腺嘌呤核苷酸的测量结果表明，苯甲酸代谢并未充分改变能量状态，无法解释所观察到的抑制现象。与这些解释一致的是，添加甘氨酸加速苯甲酰辅酶A向马尿酸盐的转化，恢复了游离辅酶A和乙酰辅酶A的水平以及糖异生和尿素生成的速率。天冬氨酸和谷氨酸水平的降低，可能是由于干扰了丙酮酸羧化酶的回补功能，很可能是苯甲酸抑制糖异生的原因。尿素循环通量降低是否也导致了糖异生的抑制（通过减少胞质溶胶中天冬氨酸向草酰乙酸的转化）还需要进一步研究。谷氨酸和乙酰辅酶A降低至NAG合成酶的Km值或以下水平，可能是所观察到的尿素生成抑制的原因。观察到尿素生成速率随NAG水平的变化而变化，表明NAG依赖性CPS - I是苯甲酸抑制尿素生成的主要位点。
