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首页> 外文期刊>Applied Microbiology >Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus
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Two Pathways for Glutamate Biosynthesis in the Syntrophic Bacterium Syntrophus aciditrophicus

机译：枯萎菌嗜酸菌中谷氨酸生物合成的两种途径

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The anaerobic metabolism of crotonate, benzoate, and cyclohexane carboxylate by Syntrophus aciditrophicus grown syntrophically with Methanospirillum hungatei provides a model to study syntrophic cooperation. Recent studies revealed that S. aciditrophicus contains Re -citrate synthase but lacks the common Si -citrate synthase. To establish whether the Re -citrate synthase is involved in glutamate synthesis via the oxidative branch of the Krebs cycle, we have used [1-~(13)C]acetate and [1-~(14)C]acetate as well as [~(13)C]bicarbonate as additional carbon sources during axenic growth of S. aciditrophicus on crotonate. Our analyses showed that labeled carbons were detected in at least 14 amino acids, indicating the global utilization of acetate and bicarbonate. The labeling patterns of alanine and aspartate verified that pyruvate and oxaloacetate were synthesized by consecutive carboxylations of acetyl coenzyme A (acetyl-CoA). The isotopomer profile and ~(13)C nuclear magnetic resonance (NMR) spectroscopy of the obtained [~(13)C]glutamate, as well as decarboxylation of [~(14)C]glutamate, revealed that this amino acid was synthesized by two pathways. Unexpectedly, only the minor route used Re -citrate synthase (30 to 40%), whereas the majority of glutamate was synthesized via the reductive carboxylation of succinate. This symmetrical intermediate could have been formed from two acetates via hydration of crotonyl-CoA to 4-hydroxybutyryl-CoA. 4-Hydroxybutyrate was detected in the medium of S. aciditrophicus when grown on crotonate, but an active hydratase could not be measured in cell extracts, and the annotated 4-hydroxybutyryl-CoA dehydratase (SYN_02445) lacks key amino acids needed to catalyze the hydration of crotonyl-CoA. Besides Clostridium kluyveri , this study reveals the second example of a microbial species to employ two pathways for glutamate synthesis.
机译：与饥饿型甲基螺旋藻一起营养生长的嗜酸性拟南芥对巴豆酸，苯甲酸酯和环己烷羧酸盐的厌氧代谢提供了研究同养协同的模型。最近的研究表明，嗜酸链球菌含有柠檬酸合酶，但缺乏常见的柠檬酸合酶。为了确定Re-柠檬酸合酶是否通过Krebs循环的氧化分支参与谷氨酸的合成，我们使用了[1-〜（13）C]乙酸酯和[1-〜（14）C]乙酸酯以及[ 〜（13）C]碳酸氢盐作为巴豆酸盐上嗜酸链霉菌的轴突生长的附加碳源。我们的分析表明，在至少14个氨基酸中检测到标记的碳，表明乙酸盐和碳酸氢盐的全球利用。丙氨酸和天冬氨酸的标记模式证明了丙酮酸和草酰乙酸是通过连续的乙酰辅酶A（乙酰辅酶A）羧化而合成的。所获得的[〜（13）C]谷氨酸的同位素结构和〜（13）C核磁共振（NMR）波谱以及[〜（14）C]谷氨酸的脱羧反应表明，该氨基酸是由两条途径。出乎意料的是，只有少数途径使用柠檬酸合酶（30％至40％），而大部分谷氨酸是通过琥珀酸酯的还原羧化反应合成的。这种对称的中间体可能是由两种乙酸酯通过巴豆酰辅酶A水合为4-羟基丁酰辅酶A形成的。在巴豆酸盐培养基上生长时，在嗜酸链霉菌的培养基中检测到4-羟基丁酸，但无法在细胞提取物中检测到活性水合酶，并且带注释的4-羟基丁酰-CoA脱水酶（SYN_02445）缺乏催化水合所需的关键氨基酸。巴豆酰辅酶A。除了克鲁维氏梭菌外，这项研究还揭示了利用两个途径合成谷氨酸的微生物物种的第二个例子。

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