Investigating the Effect of Alcohol Dehydrogenase Gene Knockout on Lipid Accumulation in Mucor circinelloides WJ11

Mucor circinelloides is an oleaginous, dimorphic zygomycete fungus species that produces appreciable levels of ethanol when grown under aerobic conditions in the presence of high glucose, indicating the fungus is a Crabtree-positive microorganism. Engineering efforts to redirect carbon flux from ethanol to lipid biosynthesis may shed light on the critical role of ethanol biosynthesis during aerobic fermentation in M. circinelloides. Therefore, in this study, the alcohol dehydrogenase gene (ADH1) of M. circinelloides WJ11 was deleted, and its effects on growth, lipid production, and fatty acid content were analyzed. Our results showed that knocking out of adh1 increment reduced the ethanol concentration by 85-90% in fermented broth, indicating that this gene is the major source of ethanol production. Parallel to these findings, the lipid and fatty acid content of the mutant was decreased, while less change in the growth of WJ11 was observed. Furthermore, a fermentation study showed the lipid and fatty acid content was restored in the mutant strain when the fermentation media was supplemented with 0.5% external ethanol, indicating the importance of alcohol dehydrogenase and its product on growth and lipid biosynthesis in M. circinelloides. To our knowledge, this is the first study to show a link between alcohol dehydrogenase and lipid production in M. circinelloides.