CARBOXYLATION OF PYRUVATE AND ACETYL COENZYME-A BY REVERSAL OF THE NA+ PUMPS OXALOACETATE DECARBOXYLASE AND METHYLMALONYL-COA DECARBOXYLASE
CARBOXYLATION OF PYRUVATE AND ACETYL COENZYME-A BY REVERSAL OF THE NA+ PUMPS OXALOACETATE DECARBOXYLASE AND METHYLMALONYL-COA DECARBOXYLASE
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DOI：
10.1021/bi00317a039
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发表时间：
1984-01-01
期刊：
影响因子：
2.9
通讯作者：
HILPERT, W
中科院分区：
文献类型：
--
作者：
DIMROTH, P;HILPERT, W
蛋白脂质体重建洗涤剂透析纯化[产气克雷伯氏菌]草酰乙酸脱羧酶和磷脂催化草酰乙酸-14 CO2交换。类似地制备的含有[Veillonella alcalescens]甲基丙二酰辅酶A脱羧酶的蛋白脂质体催化丙二酰辅酶A和14 CO2之间的交换。这些交换反应完全依赖于部分底物脱羧过程中建立的Na+离子梯度，因为在Na+载体莫能菌素的存在下没有发生交换。在将大的Na+浓度梯度Nain+ > Naout+施加到含有甲基丙二酰辅酶A脱羧酶的蛋白脂质体之后，乙酰辅酶A被羧化为丙二酰辅酶A，丙二酰辅酶A被脂肪酸合成酶捕获。在不存在Na+梯度的情况下，不发生乙酰辅酶A羧化。当草酰乙酸脱羧酶和甲基丙二酰辅酶A脱羧酶同时纳入到蛋白脂质体，转羧酶系统的构建。草酰乙酸脱羧为丙酮酸后，乙酰辅酶A羧化为丙二酰辅酶A，反之亦然。这些转羧介导的Na+电路，因为耗散的Na+梯度与莫能菌素也废除了转羧反应。用缬氨霉素或羰基氰对-（三氟甲氧基）-苯腙破坏膜电位严重降低草酰乙酸脱羧依赖性乙酰辅酶A羧化的速率，与Na+泵的产电性质雅阁。因此，这些Na+泵通过在脱羧时产生电化学Na+梯度或通过CO2固定以牺牲已经存在的Na+梯度来产生羧酸而充当可逆矢量催化剂。这种类型的CO2固定是新的，基本上不同于经典的羧化反应，需要ATP水解，以克服能量上不利的代谢过程。Na+运输和丙二酰辅酶A脱羧之间的化学计量比为2：1，在初始阶段，但降低后，Na+浓度梯度已开发的囊泡膜。
Proteoliposomes reconstituted by detergent dialysis from purified [Klebsiella aerogenes] oxaloacetate decarboxylse and phospholipids catalyzed an oxaloacetate-14CO2 exchange. Similarly prepared proteoliposomes containing [Veillonella alcalescens] methylmalonyl-CoA decarboxylase catalyzed the exchange between malonyl-CoA and 14CO2. These exchange reactions were completely dependent on the Na+ ion gradients established during decarboxylation of part of the substrates since no exchange took place in the presence of the Na+ carrier monensin. After a large Na+ concentration gradient Nain+ > Naout+ was applied to methylmalonyl-CoA decarboxylase containing proteoliposomes, acetyl-CoA was carboxylated to malonyl-CoA which was trapped with fatty acid synthetase. In the absence of a Na+ gradient, no acetyl-CoA carboxylation occurred. When oxaloacetate decarboxylase and methylmalonyl-CoA decarboxylase were simultaneously incorporated into proteoliposomes, a transcarboxylase system was constructed. Upon decarboxylation of oxaloacetate to pyruvate, acetyl-CoA was carboxylated to malonyl-CoA and vice versa. These transcarboxylations are mediated by a Na+ circuit since dissipation of the Na+ gradient with monensin also abolished the transcarboxylation reactions. Disruption of the membrane potential with valinomycin or carbonyl cyanide p-(trifluoromethoxy)-phenylhydrazone severely reduced the rate of oxaloacetate decarboxylation dependent acetyl-CoA carboxylation, in accord with the electrogenic properties of the Na+ pumps. These Na+ pumps therefore act as reversible vectorial catalysts either by creating electrochemical Na+ gradients upon decarboxylation or by CO2 fixation to yield carboxylic acids at the expense of an already existing Na+ gradient. This type of CO2 fixation is new and basically different from the classical carboxylation reactions which require ATP hydrolysis to overcome the energetically unfavorable metabolic processes. The stoichiometry between Na+ transport and malonyl-CoA decarboxylation was 2:1 in the initial phase but decreased after a Na+ concentration gradient had developed over the vesicular membrane.