The role of guanosine deaminase in ureide production in soybean nodules and in purine nucleotide catabolism of Arabidopsis

嘌呤核苷酸可以在植物中被完全分解代谢以回收营养。我们最近描述了一种来自拟南芥的酶，鸟苷脱氨酶（GSDA），参与了这一过程。GSDA催化鸟苷脱氨为黄苷，黄苷在拟南芥中仅由GSDA合成。其他来源的黄嘌呤核苷，如XMP的去磷酸化，似乎不相关。然而，在大豆中，通常认为酰脲（嘌呤碱分解产物，其充当从根瘤到芽的主要氮运输化合物）的产生通过XMP去磷酸化进行。如果这是真的，那么根瘤中酰脲的产生就不需要大豆的GSDA。(1)我们建议通过实验评估大豆根瘤中酰脲生成的途径，并评估GSDA在此过程中的作用。在拟南芥中，我们的初步实验表明，GSDA可能是至关重要的所有或至少大部分的嘌呤核苷酸catenase（GMP和AMP）。(2)通过对一系列单突变体和双突变体进行代谢物分析，绘制嘌呤核苷酸分解代谢途径图，并研究GSDA的总体重要性。GSDA突变体表现出不同的表型，如发芽率低和对长时间黑暗敏感。(3)将在嘌呤核苷酸催化剂的其他突变体的背景下对GSDA突变体进行详细的表型分析，以评价和量化观察到的缺陷。我们推测，缺陷的糖再动员可能是部分负责表型的改变。GSDA是一种新的酶，只在植物中发现。微生物具有结构相似的酶，对鸟嘌呤具有特异性。(4)通过从拟南芥中产生GSDA的晶体结构，我们希望了解这种酶的鸟苷特异性的分子基础。

Purine nucleotides can be fully catabolized in plants to recycle nutrients. We recently described an enzyme from Arabidopsis, guanosine deaminase (GSDA), involved in this process. GSDA catalyzes the deamination of guanosine to xanthosine which in Arabidopsis is exclusively synthesized by GSDA. Other sources for xanthosine, like the dephosphorylation of XMP, seem not to be relevant. In soybean however, it is generally assumed that the generation of the ureides (purine base breakdown products that serve as primary nitrogen transport compounds from the nodules to the shoot) runs via XMP dephosphorylation. If true, GSDA of soybean would not be required for ureide production in nodules. (1) We propose to experimentally evaluate the route of ureide generation in soybean nodules and assess the role of GSDA in this process. In Arabidopsis, our preliminary experiments indicated that GSDA may be of central importance for all or at least the bulk of purine nucleotide catabolism (of GMP and also AMP). (2) By performing metabolite analyses of a range of single and double mutants, a map of of the purine nucleotide catabolic pathway will be drawn and the overall importance of GSDA investigated. This will potentially result in a completely new view on purine nucleotide catabolism in vivo.Arabidopsis mutants of GSDA show distinct phenotypes like low germination and susceptibility to prolonged darkness. (3) A detailed phenotypic analysis of GSDA mutants in the context of other mutants of purine nucleotide catabolism will be performed to evaluate and quantify the observed defects. We speculate that defective sugar remobilization may be in part responsible for the phenotypic alterations.GSDA is a novel enzyme only found in plants. Microbes have structurally similar enzymes with specificity for guanine. (4) By generating a crystal structure of GSDA from Arabidopsis we would like to understand the molecular basis for the guanosine specificity of this enzyme.