Laboratory evolution and subsequent characterization of engineered Saccharomyces cerevisiae strains expressing different genes involved in cellobiose utilization

• Grant number: 18/23977-3
• Support type: Scholarships in Brazil - Scientific Initiation
• Effective date (Start): March 01, 2019
• Effective date (End): November 30, 2020
• Field of knowledge: Interdisciplinary Subjects
• Principal researcher: Andreas Karoly Gombert
• Grantee: Jennifer Alves Lanza
• Home Institution: Faculdade de Engenharia de Alimentos (FEA). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil

Abstract

Cellobiose is a disaccharide present in the context of a second-generation (2G) biorefinery, which employs lignocelullosic raw materials for the production of bio-based fuels and chemicals. The fuel ethanol industry currently uses the yeast Saccharomyces cerevisiae as the microbial platform for the production of ethanol via fermentation. However, S. cerevisiae is not able to naturally metabolize cellobiose, an intermediate product that accumulates during the enzymatic hydrolysis of cellulosic materials, reducing its rate and efficiency. Introduction of an intracellular cellobiose assimilation pathway into S. cerevisiae has been proposed as a good alternative to circumvent this and some other relevant issues regarding 2G ethanol production. In a previous work, six S. cerevisiae strains expressing different fungal genes involved in cellobiose utilization were engineered. These genes encode a cellodextrin transporter and six different intracellular beta-glucosidases, five of them being novel enzymes not described in the literature so far. Although these strains have not been studied in detail, it has been observed that their growth in cellobiose-containing media is rather slow. Thus, in this project we intend to improve the kinetic perfomances of the engineered strains by laboratory evolution, a popular methodology applied for improving industrially relevant traits in yeast. Subsequently, the resulting evolved and parental phenotypes will be characterized by molecular and physiological studies in shake flasks with cellobiose as the sole carbon and energy source.