Engineered yeast for enzymatic hydrolysis of laminarin from brown macroalgae

Abstract Increasing demands for fuel production have led to the identification of brown macroalgae as an alternative feedstock for biofuel production. However, the conversion of macroalgae to biofuel using yeast, such as Saccharomyces cerevisiae, requires the production of enzymes capable of hydrolysing macroalgal carbohydrates. In this study, three synthetic laminarinase-encoding genes from Rasamsonia emersonii (Relam1), Trichoderma viride (Tvlam1) and Trichoderma reesei (Trlam1), as well as a native laminarinase-like gene from S. cerevisiae (Sclam1) were cloned and expressed in the laboratory S. cerevisiae Y294 strain. Supernatant containing the Relam1 enzyme displayed the highest extracellular activity at 45 °C (0.2 U/mL), while supernatant from the strain co-expressing the Relam1 and Tvlam1 genes showed the highest extracellular activity at 37 °C (0.09 U/mL). The Relam1 and Tvlam1 laminarinases displayed optimum enzymatic activity at temperatures of 45 °C and 37 °C, respectively, and Relam1 retained more than 89% activity after 96 h at 45 °C. Large-scale fermentation of Ecklonia maxima macroalgae with S. cerevisiae Y294[Re/Tvlam1] in a benchtop bioreactor achieved 43% of the theoretic ethanol yield and 66% carbon conversion from laminarin. The successful expression of laminarinase-encoding genes in S. cerevisiae is an important milestone in exploiting brown macroalgae for bioethanol production via consolidated bioprocessing.