Model-Driven Elucidation of Lactose and Galactose Metabolism via Oxidoreductive Pathway inSungouiella intermediafor Cell Factory Applications

Abstract

Converting industrial side streams into value-added chemicals using microbial cell factories is of increasing interest, as such processes offer solutions to reduce waste and production costs. However, developing new, efficient cell factories for precision fermentation remains challenging due to limited knowledge about their metabolic capabilities. Here, we investigate the lactose and galactose metabolism of the non-conventional yeast Sungouiella intermedia (formerly Candida intermedia), using knowledge-matching of high-quality genome-scale metabolic model (GEM) with extensive experimental analysis and determine its potential as a future cell factory on lactose-rich industrial side-streams. We show that this yeast possesses the conserved Leloir pathway as well as an oxidoreductive galactose catabolic route. Contextualization of RNAseq data into Sint-GEM highlights the regulatory mechanisms on the oxidoreductive pathway and how this pathway can enable adaptation to diverse environments. Model simulations, together with experimental data from continuous and batch bioreactors, indicate that S. intermedia uses upstream enzymes of the oxidoreductive pathway, in a condition-dependent manner, and produce the sugar alcohol galactitol as a carbon overflow metabolite, coupled to redox co-factor balancing during both lactose and galactose growth. Furthermore, the new metabolic insights facilitated the development of an improved bioprocess design, where an engineered S. intermedia strain could achieve galactitol yields of >90% of the theoretical maximum at improved production rates using the industrial side-stream cheese whey permeate as feedstock. Additional strain engineering resulted in galactitol-to-tagatose conversion, proving the versatility of the future production host. Overall, this work sheds new light on the intrinsic interplay between parallel metabolic pathways that shape the lactose and galactose catabolism in S. intermedia. It also demonstrates how a GEM combined with experimental analysis can work in synergy to fast-forward metabolic characterization and development of new, non-conventional yeast cell factories.