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叶酸。人类需要的叶酸主要来自植物性食物。叶酸分子有三个部分--蝶啶、对氨基苯甲酸(PABA)和谷氨酸--通常还有一条多谷氨酸的尾巴。本研究有三个目的。(A)通过转基因提高番茄果实中的叶酸水平，使其同时产生蝶啶和PABA(从以前的工作中可以清楚地看到，仅过量生产蝶啶的水果含有少量的叶酸，但已经用完了PABA)，并分析了基因工程水果中叶酸合成基因的表达情况。(B)将基因组学和生物化学结合起来，以确定和确定回收叶酸分解产物所涉及的酶和基因的特征，因为在化学或光化学分解比率较高的情况下，这种回收似乎对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伽马-谷氨酰水解酶活性，因为这种酶攻击多谷氨酸叶酸，并可能实际上针对它们进行破坏，因为脱谷氨酸叶酸更容易在体内分解。广泛的影响：缺乏维生素叶酸是一个全球健康问题，导致可预防的出生缺陷和贫血的巨大负担，并增加血管疾病和癌症的风险。在植物或其他食物有机体中进行叶酸代谢工程(生物强化)是纠正膳食叶酸缺乏的一种可行方法，在几个国家正在进行中。这项研究将推动这一国际研究和开发努力，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，并提高通过生物强化改善叶酸营养的潜力。这一推广活动将包括关于测量食物中叶酸的演讲和简单的实验室练习，这将展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.