Identification of alanine dehydrogenase and its role in mixed secretion of ammonium and alanine by pea bacteroids
Identification of alanine dehydrogenase and its role in mixed secretion of ammonium and alanine by pea bacteroids
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DOI：
10.1046/j.1365-2958.2000.01884.x
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发表时间：
2000-04-01
影响因子：
3.6
通讯作者：
Poole, PS
中科院分区：
文献类型：
--
作者：
Allaway, D;Lodwig, EM;Poole, PS
根瘤菌-豆科植物共生体的固氮作用具有重要的生态和农业意义，是生物圈氮素的主要来源。根据N-15标记研究，人们普遍认为铵是类杆菌固氮的唯一分泌产物，植物负责将其同化为氨基酸。然而，最近的一项N-15标记研究表明，大豆类杆菌只分泌丙氨酸，这一范式受到了挑战。然而，仅从分离的类菌体的体外N-15标记研究中评估了氮分泌。我们发现，铵和丙氨酸分泌豌豆类杆菌。它们之间的体外分配将取决于系统是开放的还是封闭的，以及铵浓度和类菌体密度。为了克服这些局限性，我们确定和突变的基因丙氨酸脱氢酶（aldA），并证明AldA是丙氨酸合成的主要途径，在分离的类菌体。aldA突变体的拟杆菌固定氮，但仅以显著速率分泌铵，导致总氮分泌较低。用aldA突变体接种的豌豆是绿色和健康的，表明类杆菌的铵分泌可以为植物生长提供足够的氮。然而，与接种野生型的那些相比，接种突变体的植物的生物量减少。标记和植物生长研究表明，丙氨酸的合成和分泌有助于N-2固定的效率，因此生物量积累。
N-2-fixation by Rhizobium-legume symbionts is of major ecological and agricultural importance, responsible for producing a substantial fraction of the biosphere's nitrogen. On the basis of N-15-labelling studies, it had been generally accepted that ammonium is the sole secretion product of N-2-fixation by the bacteroid and that the plant is responsible for assimilating it into amino acids. However, this paradigm has been challenged in a recent N-15-labelling study showing that soybean bacteroids only secrete alanine. Hitherto, nitrogen secretion has only been assessed from in vitro N-15-labelling studies of isolated bacteroids. We show that both ammonium and alanine are secreted by pea bacteroids. The in vitro partitioning between them will depend on whether the system is open or closed, as well as the ammonium concentration and bacteroid density. To overcome these limitations we identified and mutated the gene for alanine dehydrogenase (aldA) and demonstrate that AldA is the primary route for alanine synthesis in isolated bacteroids. Bacteroids of the aldA mutant fix nitrogen but only secrete ammonium at a significant rate, resulting in lower total nitrogen secretion. Peas inoculated with the aldA mutant are green and healthy, demonstrating that ammonium secretion by bacteroids can provide sufficient nitrogen for plant growth. However, plants inoculated with the mutant are reduced in biomass compared with those inoculated with the wild type. The labelling and plant growth studies suggest that alanine synthesis and secretion contributes to the efficiency of N-2-fixation and therefore biomass accumulation.