Engineering Enzymatic Pathways in the Gut Microbiome to Control Dietary Isoflavone Metabolism

Abstract

**Please note that the full text is embargoed until 5/17/2024** ABSTRACT: Observing how plasmids persist within the gut can help us understand how newly introduced genes are transmitted within the gut microbiome. I sought to determine the persistence of genetic constructs introduced to the murine gut and then develop our own genetic system to be delivered to the gut to engineer the microbiome in its ability to convert certain dietary isoflavones. In my initial experiments, mice were given bacteria containing a genetically engineered plasmid and were examined for the persistence of the plasmid in the gut. I found long-term persistence of the plasmid in the gut for up to 2 months when administering antibiotics during and following 1 week dosing of the mice with bacteria carrying the plasmid. The use of higher concentrations of carrier bacteria influenced the short-term abundance of the plasmid-carrying bacteria in the gut. I developed a bacterial expression system for the production of enzymes capable of metabolizing dietary isoflavones into products known to serve as estrogen receptor agonists, specifically the metabolite equol, with potential health benefits including protecting against cardiovascular disease and ischemic stroke. In developing this and proving the production of equol in vitro, I moved to using this system in murine studies. In vivo examination identified that the probiotic I developed could impart increased presence of serum equol concentrations with dietary soy. Finally, I examined the impact of dietary isoflavones on relative abundance and diversity of bacteria in the murine gut.