Coupling between d-3-phosphoglycerate dehydrogenase and d-2-hydroxyglutarate dehydrogenase drives bacterial l-serine synthesis

Significance d-3-Phosphoglycerate dehydrogenase (SerA) is a key enzyme in l-serine biosynthesis. It couples the dehydrogenation of d-3-phosphoglycerate to 3-phosphohydroxypyruvate and the reduction of 2-ketoglutarate to d-2-hydroxyglutarate (d-2-HG). This provides an example of how nonenergetically favorable and energetically favorable reactions are linked together to allow metabolic processes to proceed. d-2-HG is often considered as an abnormal metabolite produced by several enzymes with “promiscuous” activities. Our findings offer insights into how an enzymatic reaction that was considered promiscuous or accidental plays a key role in metabolism. We have identified a bacterial d-2-hydroxyglutarate dehydrogenase (D2HGDH), which converts d-2-HG produced during l-serine biosynthesis back to 2-ketoglutarate. d-2-HG is a normal metabolite that is simultaneously produced and catabolized without accumulation in bacterial metabolism. l-Serine biosynthesis, a crucial metabolic process in most domains of life, is initiated by d-3-phosphoglycerate (d-3-PG) dehydrogenation, a thermodynamically unfavorable reaction catalyzed by d-3-PG dehydrogenase (SerA). d-2-Hydroxyglutarate (d-2-HG) is traditionally viewed as an abnormal metabolite associated with cancer and neurometabolic disorders. Here, we reveal that bacterial anabolism and catabolism of d-2-HG are involved in l-serine biosynthesis in Pseudomonas stutzeri A1501 and Pseudomonas aeruginosa PAO1. SerA catalyzes the stereospecific reduction of 2-ketoglutarate (2-KG) to d-2-HG, responsible for the major production of d-2-HG in vivo. SerA combines the energetically favorable reaction of d-2-HG production to overcome the thermodynamic barrier of d-3-PG dehydrogenation. We identified a bacterial d-2-HG dehydrogenase (D2HGDH), a flavin adenine dinucleotide (FAD)-dependent enzyme, that converts d-2-HG back to 2-KG. Electron transfer flavoprotein (ETF) and ETF-ubiquinone oxidoreductase (ETFQO) are also essential in d-2-HG metabolism through their capacity to transfer electrons from D2HGDH. Furthermore, while the mutant with D2HGDH deletion displayed decreased growth, the defect was rescued by adding l-serine, suggesting that the D2HGDH is functionally tied to l-serine synthesis. Substantial flux flows through d-2-HG, being produced by SerA and removed by D2HGDH, ETF, and ETFQO, maintaining d-2-HG homeostasis. Overall, our results uncover that d-2-HG–mediated coupling between SerA and D2HGDH drives bacterial l-serine synthesis.

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