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.
