Lee Hyo, Park Sangkyu, Lee Saet Buyl, Song Jaeeun, Kim Tae-Hwan, Kim Beom-Gi
Metabolic Engineering Division, National Institute of Agricultural Sciences, Rural Development Administration, Jeonju, Republic of Korea.
Department of Animal Science, Institute of Agricultural Science and Technology, College of Agriculture and Life Science, Chonnam National University, Gwangju, Republic of Korea.
Front Plant Sci. 2024 Oct 18;15:1464877. doi: 10.3389/fpls.2024.1464877. eCollection 2024.
The flavonoid diosmin (diosmetin 7--rutinoside) is used as a therapeutic agent for disorders of the blood vessels such as hemorrhoids and varicose veins. Diosmin is commercially produced using semi-synthetic methods involving the oxidation of hesperidin, the most abundant flavonoid in citrus fruits. However, this method produces byproducts that are toxic to the environment, and new sustainable methods to produce diosmin are required. Here, we used a synthetic biology approach to produce diosmin without generating toxic byproducts through reconstitution of the diosmin biosynthetic pathway in . We first established that leaves co-infiltrated with all seven genes in the flavonoid biosynthesis pathway produced high levels of luteolin, a precursor of diosmetin. We then compared the activity of modification enzymes such as methyltransferases, glucosyltransferases, and rhamnosyltransferases in and and selected genes encoding enzymes with the highest activity for producing diosmetin, diosmetin 7--glucoside, and diosmin, respectively. Finally, we reconstructed the entire diosmin biosynthetic pathway using three constructs containing ten genes encoding enzymes in this pathway, from phenylalanine ammonia lyase to rhamnosyltransferase. leaves transiently co-expressing all these genes yielded 37.7 µg diosmin per gram fresh weight. To our knowledge, this is the first report of diosmin production in a heterologous plant system without the supply of a precursor. Successful production of diosmin in opens new avenues for producing other commercially important flavonoids using similar platforms.
类黄酮地奥司明（香叶木素7 - 芸香糖苷）被用作治疗血管疾病（如痔疮和静脉曲张）的治疗剂。地奥司明是通过半合成方法商业生产的，该方法涉及柑橘类水果中最丰富的类黄酮橙皮苷的氧化。然而，这种方法会产生对环境有毒的副产物，因此需要新的可持续生产地奥司明的方法。在这里，我们使用合成生物学方法，通过在植物中重建地奥司明生物合成途径来生产地奥司明，而不产生有毒副产物。我们首先确定，与类黄酮生物合成途径中的所有七个基因共浸润的植物叶片产生了高水平的木犀草素，这是香叶木素的前体。然后，我们比较了植物和植物中甲基转移酶、葡萄糖基转移酶和鼠李糖基转移酶等修饰酶的活性，并分别选择了编码产生香叶木素、香叶木素7 - 葡萄糖苷和地奥司明活性最高的酶的基因。最后，我们使用三个构建体重建了整个地奥司明生物合成途径，这些构建体包含该途径中从苯丙氨酸解氨酶到鼠李糖基转移酶的十个编码酶的基因。瞬时共表达所有这些基因的植物叶片每克鲜重产生37.7微克地奥司明。据我们所知，这是在不供应前体的异源植物系统中生产地奥司明的首次报道。在地奥司明在植物中的成功生产为使用类似平台生产其他商业上重要的类黄酮开辟了新途径。
