叶酸调控拟南芥谷氨酰胺合成酶活性的分子机制

叶酸属水溶性B族维生素（B9），是生物体生长发育的重要辅因子。近来的研究表明，叶酸参与包括氮代谢在内的多种生物学过程，但其详细分子机制尚不清楚。为此，我们课题组在前期工作中进行了叶酸探针制备并利用点击化学（click chemistry）和定量蛋白质组学技术鉴定了拟南芥叶酸互作蛋白，发现拟南芥谷氨酰胺合成酶蛋白（AtGLNs）与叶酸探针互作，微量热泳动实验（microscale thermophoresis，MST）证明了AtGLN1;4蛋白与叶酸分子的结合作用。在上述工作基础上，本项目旨在利用结构生物学技术进一步明确叶酸分子与AtGLNs蛋白的结合模式与结合位点，利用酶促反应动力学实验阐明叶酸分子结合对AtGLNs酶学活性的影响，探索植物体内叶酸-AtGLNs互作对植物氮代谢的影响，从而解析叶酸参与植物氮代谢调控的分子机制，为进一步深入理解植物氮代谢调控机理提供新的理论基础。

Folates, a class of vitamin B, are essential cofacotors for growth and development of almost all organisms. Recent study has shown that folates take part in several metabolic pathways in plants including nitrogen (N) metabolism; however, the molecular mechanisms remain completely unknown. To uncover this, we synthesized a folate probe and utilized “click chemistry” in combination with quantitative proteomics to identify the folate-interacting proteins in Arabidopsis plants. Glutamine synthetases (AtGLNs) were found to have high affinity with the folate probe. Further, the interaction of folates with AtGLN1;4 was validated by microscale thermophoresis (MST). In this study, we aim to explore the acting mode and binding site of AtGLNs with folates via structural biology, to investigate the impact of the interaction on enzymatic activities of AtGLNs by kinetics of enzyme-catalyzed reactions and elucidate the effect of folates-AtGLNs interaction on plant nitrogen metabolism. These results will help understand the molecular mechanism underlying folates’ participation in plant nitrogen metabolism from a new angle.