Pyruvate is synthesized by two pathways in pea bacteroids with different efficiencies for nitrogen fixation.
Pyruvate is synthesized by two pathways in pea bacteroids with different efficiencies for nitrogen fixation.
复制标题
丙酮酸在豌豆类菌中通过两种途径合成，固氮效率不同。
DOI：
10.1128/jb.00294-10
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发表时间：
2010
影响因子：
3.2
通讯作者：
Mulley G
中科院分区：
文献类型：
--
作者：
Mulley G
豆科类杆菌中的固氮是由二羧酸的代谢提供能量的，这需要它们氧化成草酰乙酸和丙酮酸。在苜蓿拟杆菌中，丙酮酸的产生需要NAD+苹果酸酶（Dme），而不需要NADP+苹果酸酶（Tme）。然而，我们发现豆科根瘤菌有两条途径从二羧酸的丙酮酸形成由Dme催化和由磷酸烯醇式丙酮酸（PEP）羧激酶（PckA）和丙酮酸激酶（PykA）的联合活动。这两种途径都能使N2固定，但PckA/PykA途径仅支持Dme的60%的N2固定。dme和pckA/pykA双突变体不能固定N2。此外，dme pykAdouble突变体不能在二羧酸盐上生长，表明它们是从正常表达的二羧酸盐生产丙酮酸盐的唯一途径。pckA在苜蓿类杆菌中不表达，导致丙酮酸形成和N2固定对Dme的专性需求。当PckA从alfalfadmebacteroids中的constitutivenptII启动子表达时，乙炔以野生型速率的30%减少，尽管该水平不足以防止氮饥饿。Dme具有N-末端、苹果酸酶（Me）和C-末端磷酸转乙酰酶（Pta）结构域。删除Pta结构域增加了峰值乙炔还原率在4周龄豌豆植物的野生型率的140至150%，这是伴随着增加结节质量。感染Pta缺失突变体的植物没有增加干重，表明在整个生长过程中固氮没有持续的变化。这表明类杆菌中丙酮酸合成、固氮和植物生长之间存在复杂的关系。
Nitrogen fixation in legume bacteroids is energized by the metabolism of dicarboxylic acids, which requires their oxidation to both oxaloacetate and pyruvate. In alfalfa bacteroids, production of pyruvate requires NAD+malic enzyme (Dme) but not NADP+malic enzyme (Tme). However, we show thatRhizobium leguminosarumhas two pathways for pyruvate formation from dicarboxylates catalyzed by Dme and by the combined activities of phosphoenolpyruvate (PEP) carboxykinase (PckA) and pyruvate kinase (PykA). Both pathways enable N2fixation, but the PckA/PykA pathway supports N2fixation at only 60% of that for Dme. Double mutants ofdmeandpckA/pykAdid not fix N2. Furthermore,dme pykAdouble mutants did not grow on dicarboxylates, showing that they are the only pathways for the production of pyruvate from dicarboxylates normally expressed. PckA is not expressed in alfalfa bacteroids, resulting in an obligate requirement for Dme for pyruvate formation and N2fixation. When PckA was expressed from a constitutivenptIIpromoter in alfalfadmebacteroids, acetylene was reduced at 30% of the wild-type rate, although this level was insufficient to prevent nitrogen starvation. Dme has N-terminal, malic enzyme (Me), and C-terminal phosphotransacetylase (Pta) domains. Deleting the Pta domain increased the peak acetylene reduction rate in 4-week-old pea plants to 140 to 150% of the wild-type rate, and this was accompanied by increased nodule mass. Plants infected with Pta deletion mutants did not have increased dry weight, demonstrating that there is not a sustained change in nitrogen fixation throughout growth. This indicates a complex relationship between pyruvate synthesis in bacteroids, nitrogen fixation, and plant growth.