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首页> 外文OA文献 >The mechanisms of reductive carboxylation reactions. Carbon dioxide or bicarbonate as substrate of nicotinamide-adenine dinucleotide phosphate-linked isocitrate dehydrogenase and `malic' enzyme
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The mechanisms of reductive carboxylation reactions. Carbon dioxide or bicarbonate as substrate of nicotinamide-adenine dinucleotide phosphate-linked isocitrate dehydrogenase and `malic' enzyme

机译：还原性羧化反应的机理。二氧化碳或碳酸氢盐作为烟酰胺-腺嘌呤二核苷酸磷酸连接的异柠檬酸脱氢酶和“苹果酸”酶的底物

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1. A simple kinetic method was devised to show whether dissolved CO2 or HCO3– ion is the substrate in enzyme-catalysed carboxylation reactions. 2. The time-course of the reductive carboxylation of 2-oxoglutarate by NADPH, catalysed by isocitrate dehydrogenase, was studied by a sensitive fluorimetric method at pH7·3 and pH6·4, with large concentrations of substrate and coenzyme and small carbon dioxide concentrations. 3. Reaction was initiated by the addition of carbon dioxide in one of three forms: (i) as the dissolved gas in equilibrium with bicarbonate; (ii) as unbuffered bicarbonate solution; (iii) as the gas or as an unbuffered solution of the gas in water. Different progress curves were obtained in the three cases. 4. The results show that dissolved CO2 is the primary substrate of the enzyme, and that HCO3– ion is at best a very poor substrate. The progress curves are in quantitative agreement with this conclusion and with the known rates of the reversible hydration of CO2 under the conditions of the experiments. The effects of carbonic anhydrase confirm the conclusions. 5. Similar experiments on the reductive carboxylation of pyruvate catalysed by the `malic' enzyme show that dissolved CO2 is the primary substrate of this enzyme also. 6. The results are discussed in relation to the mechanisms of these enzymes, and the effects of pH on the reactions. 7. The advantages of the method and its possible applications to other enzymes involved in carbon dioxide metabolism are discussed.
机译：1.设计了一种简单的动力学方法，以显示在酶催化的羧化反应中底物是溶解的CO2还是HCO3-。 2.采用敏感的荧光法，在pH7·3和pH6·4的条件下，采用底物和辅酶浓度高，二氧化碳浓度低的方法，研究了异柠檬酸脱氢酶催化NADPH催化2-氧戊二酸酯还原羧化的时程。 。 3.通过加入以下三种形式之一的二氧化碳来引发反应：（i）与碳酸氢盐平衡的溶解气体； （ii）作为无缓冲碳酸氢盐溶液； （iii）作为气体或气体在水中的无缓冲溶液。在这三种情况下获得了不同的进度曲线。 4.结果表明，溶解的CO2是酶的主要底物，而HCO3-离子充其量是非常差的底物。进度曲线与该结论以及实验条件下已知的CO2可逆水合作用速率定量一致。碳酸酐酶的作用证实了这一结论。 5.“苹果酸”酶催化丙酮酸还原羧化的类似实验表明，溶解的CO2也是该酶的主要底物。 6.讨论了有关这些酶的机理以及pH对反应的影响的结果。 7.讨论了该方法的优势及其在二氧化碳代谢中涉及的其他酶的可能应用。

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