Engineering Saccharomyces cerevisiae for the production of sophorolipid biosurfactants

Biosurfactants offer a renewable alternative to petrochemical surfactants, helping mitigate the environmental burdens associated with conventional production processes. Among glycolipid biosurfactants sophorolipids represent a particularly promising class due to their diverse structural congeners and tunable physicochemical properties. Sophorolipids, composed of a sophorose moiety linked to a hydroxylated fatty acid, are typically produced by the yeast Starmerella bombicola . Although the sophorolipid biosynthetic pathway in S. bombicola is well characterized and amenable to genetic modification, some challenges persist motivating the exploration of also heterologous production pathways. While individual sophorolipid pathway genes have been expressed in Saccharomyces cerevisiae , complete heterologous biosynthesis of sophorolipids in S. cerevisiae has not previously been achieved. Heterologous biosynthesis of sophorolipid biosurfactants was achieved through metabolic engineering of S. cerevisiae. The engineered strain produced at least 16 structurally distinct sophorolipid congeners, varying in fatty acid chain length, degree of glycosylation, and position of the sugar moiety. Unexpectedly, UGTB1 alone was sufficient to catalyze sophorolipid formation, revealing a previously unrecognized capability of this enzyme in the heterologous host. Furthermore, ω/ω-1-hydroxylation of fatty acids was identified as a major bottleneck in the pathway, whereas downstream glycosylation reactions proceeded efficiently reaching up to 82.9 ± 1.9 mg/L total glycolipid titer. This work provides proof-of-concept for producing sophorolipids in a non-native host organism and validates the functionality of the sophorolipid biosynthetic pathway in S. cerevisiae . The findings establish a foundation for expanding glycolipid structural diversity through synthetic biology and support the development of sustainable biosurfactant production platforms.