Improving cellulosic ethanol production by an engineered yeast consortium displaying a pentafunctional mini-cellulosome.

As a traditional ethanol-producing microorganism, Saccharomyces cerevisiae is an ideal host for consolidated bioprocessing. However, when overloaded cellulase genes are expressed in yeast, the metabolic burden on cells may greatly affect cell growth and cellulosic ethanol production. In this study, we developed a yeast consortium system that secretes and assembles five types of cellulases on the yeast cell surface to improve cellulosic ethanol production. This system involves one display strain, which provides the scaffoldin on the surface and several secretion strains that secrete each cellulase. The secreted dockerin-containing enzymes, cellobiohydrolase (CBH), endoglucanase (EG), β-glucosidase (BGL), cellobiose dehydrogenase (CDH), and lytic polysaccharide monooxygenase (LPMO), were randomly assembled to the scaffoldin to generate a pentafunctional mini-cellulosome via cohesion-dockerin interactions. The developed system relieved the metabolic burden placed on the engineered single yeast strain and leveraged the innate metabolic potential of each host. In addition, the enzymes in the consortium acted synergistically and efficiently boosted cellulose degradation and ethanol production. When compared with the conventional system, this consortium system increased the ethanol titers from 2.66 to 4.11 g/l with phosphoric acid swollen cellulose (PASC) as the substrate, an improvement of 55%. With Avicel as the substrate, ethanol titers increased from 1.57 to 3.24 g/l, representing an enhancement of 106%.