Diversification of rhamnolipids/glycolipids in heterologous lipogenic yeast hosts using synthetic biology

糖脂（GLs）是由多种生物，特别是酵母和细菌产生的。最突出的GLs是鼠李糖脂（RLs），存在于多种RL同系物和同源物中。虽然它们的确切生理功能尚不清楚，但rl被认为是在应对不利环境条件时产生的防御或生存机制。RLs被认为是致病微生物如铜绿假单胞菌的毒力因子，因为它们有助于人类和植物生物表面的定植。尽管如此，RLs具有有趣的抗菌、抗寄生虫、抗癌、免疫调节、抗粘附以及表面活性，使它们能够在医学、农业、环境和工业中找到几种应用。值得注意的是，这些活动随着RL结构的变化而变化，从而产生了生物和物理活动的多样性。生物和物理活动的丰富性激发了对具有潜在新颖或更有趣活动的新RLs的研究。尽管如此，在过去的几十年里，新的RL的发现已经大大放慢了速度，因为它依赖于从环境样本中分离出来的新RL生产者的传统筛选。这导致产生相同rl分子的相同rl生产者的重复鉴定。RLs商业化面临的另一个问题是其生产成本高昂，这主要是由于其最佳生产者的致病性，如铜绿假单胞菌（P. aeruginosa）是一种机会性人类病原体。rl生产者的这种致病性需要昂贵的生物控制措施，这增加了rl的净成本。在非致病性微生物宿主中过量生产结构多样化的RLs的目标下，我们为该项目设定了短期目标，以开发基于基因组挖掘的异源生产非致病性脂质体酵母宿主多种RLs的概念验证策略。这包括生物合成RLs的合成代谢回路的设计，将遗传结构构建并整合到脂性耶氏菌中，最后测试和鉴定所产生的RLs。这种设计-建造-测试的合成生物学循环将反复重复，直到产生所需的RL同系物或同源物。在随后的发现资助中，我们将使用这些策略以令人满意的产量和低成本生产有趣的RL同源物或同源物。该研究项目最终将以低成本生产出多种与有趣的生物和物理活动相关的设计RL分子，这些分子对许多治疗、环境和工业应用都很有用。它还将产生几篇原创研究文章，申请专利，培养本科生和研究生水平的高素质专业人员，他们将为加拿大不断上升的化学合成生物学领域做出称职的贡献。

Glycolipids (GLs) are produced by diverse organisms, particularly by yeasts and bacteria. The most prominent GLs are rhamnolipids (RLs) that exist in a diversity of RL congeners and homologs. Although their exact physiological functions are unknown, RLs are thought to be produced as defense or survival mechanisms in response to unfavorable environmental conditions. RLs are considered virulence factors of pathogenic microorganisms like Pseudomonas aeruginosa as they help in the colonization of biotic surfaces of humans and plants. Nonetheless, RLs have interesting antimicrobial, antiparasitic, anticancer, immunomodulatory, anti-adherent as well as surface activities allowing them to find several medical, agricultural, environmental, and industrial applications. It is noteworthy that these activities are vary with variations in RL structures creating thus a diversity of biological and physical activities. This richness in biological and physical activities incited the search for new RLs with potentially novel or more interesting activities. Nonetheless, the discovery of new RLs has slowed down drastically in the last decades because it relied on classical screening of new RL producers isolated from environmental samples. This resulted in the redundant identifications of the same RL-producers that are producing the same RLs molecules. Another problem facing RLs commercialization is their costly production arising mainly from the pathogenic nature of their best producers, like P. aeruginosa which is an opportunistic human pathogen. Such pathogenic nature of RL-producers requires costly biocontainment measures which adds to the net cost of RLs. Under the goal to overproduce structurally diverse RLs in non-pathogenic microbial hosts, we set our short-term objective for program to develop proof-of-concept strategies for the genome-mining based heterologous production of diverse RLs in the non-pathogenic lipogenic yeast host, Yarrowia lipolytica. This involves the design of synthetic metabolic circuits biosynthesizing RLs, building and integrating genetic constructs into Yarrowia lipolytica, and finally testing and identification of produced RLs. This design-build-test synthetic biology cycles will be iteratively repeated until the desired RL congener or homolog is produced. In subsequent discovery grants, we will use these strategies to overproduce interesting RL congener or homologs at satisfactory yields and at low cost. This research program will ultimately result in the low-cost production of a diversity of designer RL molecules associated with interesting biological and physical activities that are useful for many therapeutic, environmental, and industrial applications. It will also result in the production of several original research articles, filing of patents and training of highly qualified professionals at undergraduate and graduate levels who will competently contribute to the rising chemical synthetic biology domain in Canada.