Folate Synthesis, Turnover, and Engineering in Plants

叶酸是所有生物体中一碳转移反应的重要辅助因子，但只有植物和微生物才能产生。人类所需的叶酸主要来自植物性食物。叶酸分子由三部分组成——蝶啶、对氨基苯甲酸酯（pABA）和谷氨酸，通常还有一个聚谷氨酸尾部。这项研究有三个目的。(a)通过设计番茄果实过量生产翼啶和pABA来提高番茄果实中的叶酸水平（从先前的工作中可以清楚地看出，仅过量生产翼啶的水果叶酸含量适度丰富，但pABA已经耗尽），并对工程水果中叶酸合成基因的表达进行分析。(b)将基因组学和生物化学结合起来，确定和鉴定参与叶酸分解产物再循环的酶和基因，因为这种再循环似乎对在化学或光化学分解率很高的情况下维持植物组织中的叶酸水平至关重要。(c)探索在番茄果实中设计较低的γ -谷氨酰水解酶活性，因为这种酶攻击多谷氨酰叶酸，并且可能实际上针对它们进行破坏，因为去谷氨酰叶酸在体内更容易分解。更广泛的影响：维生素叶酸缺乏是一个全球性的健康问题，它会导致可预防的出生缺陷和贫血的巨大负担，并增加患血管疾病和癌症的风险。植物或其他食物生物中叶酸的代谢工程（生物强化）是纠正膳食叶酸缺乏的可行方法，在一些国家正在进行中。这项研究将促进国际研究和发展的努力，并培养代谢生物化学和工程方面的博士生和博士后。此外，它还将扩大到初中和高中，以提高人们对叶酸营养的认识，以及通过生物强化来改善叶酸的潜力。这次拓展将包括演讲和简单的实验室练习，测量食物中的叶酸，这将显示水果和蔬菜的重要性。

Folates are vital cofactors for one-carbon transfer reactions in all organisms, but are made only by plants and microorganisms. The folate that humans need comes mainly from plant foods. Folate molecules have three parts - pteridine, p-aminobenzoate (pABA), and glutamate - and usually also have a polyglutamate tail. This research has three aims. (a) To raise folate levels in tomato fruit by engineering them overproduce both pteridine and pABA (it being clear from previous work that fruit overproducing pteridine alone are modestly enriched in folate, but have run out of pABA) and to profile the expression of folate synthesis genes in engineered fruit. (b) To combine genomics and biochemistry to identify and characterize enzymes and genes involved in recycling folate breakdown products, since this recycling appears to be crucial to maintaining folate levels in plant tissues in the face of high rates of chemical or photochemical breakdown. (c) To explore engineering lower gamma-glutamyl hydrolase activity in tomato fruit, since this enzyme attacks polyglutamylated folates and may in effect target them for destruction since deglutamylated folates are more prone to breakdown in vivo.Broader Impact: Deficiency of the vitamin folate is a global health problem that results in a huge burden of preventable birth defects and anemia, and increased risks of vascular disease and cancer. Metabolic engineering of folates in plants or other food organisms (biofortification) is a viable way to correct dietary folate deficiency and is in progress in several countries. This research will advance this international research and development effort, and train PhD students and postdoctorals in metabolic biochemistry and engineering. Furthermore, it will incorporate an outreach to middle and high schools to heighten awareness of folate nutrition and the potential for improving it by biofortification. This outreach will incorporate presentations and simple lab exercises on measuring folate in foods, which will show the importance of fruits and vegetables.