Biology subjects

Geijer, C.

Publications and source records attributed to Geijer, C..

3 recordsLinked to original sources

bioRxiv2024

Model-Driven Elucidation of Lactose and Galactose Metabolism via Oxidoreductive Pathway in Sungouiella intermedia for Cell Factory Applications

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. HighlightsO_LIAn oxidoreductive pathway functions in concert with the Leloir pathway for galactose catabolism. C_LIO_LIGEM predicts that galactitol secretion enables efficient carbon overflow metabolism and maintains redox balance. C_LIO_LIKnowledge-matching of GEM with experimental results highlights cell factory potential. C_LIO_LIHigh galactitol yields and proof-of-concept tagatose production using whey permeate as feedstock. C_LI

bioengineering↗

bioRxiv2023

Regulation of lactose and galactose growth: Insights from a unique metabolic gene cluster in Candida intermedia

Lactose assimilation is a relatively rare trait in yeasts, and Kluyveromyces yeast species have long served as model organisms for studying lactose metabolism. Meanwhile, the metabolic strategies of most other lactose-assimilating yeasts remain unknown. In this work, we have elucidated the genetic determinants of the superior lactose-growing yeast Candida intermedia. Through genomic and transcriptomic analyses and deletion mutant phenotyping, we identified three interdependent gene clusters responsible for the metabolism of lactose and its hydrolysis product galactose: the conserved LAC cluster (LAC12, LAC4) for lactose uptake and hydrolysis, the conserved GAL cluster (GAL1, GAL7, GAL10) for galactose catabolism, and a unique "GALLAC" cluster. This novel GALLAC cluster, which has evolved through gene duplication and divergence, proved indispensable for C. intermedias growth on lactose and galactose. The cluster contains the transcriptional activator gene LAC9, second copies of GAL1 and GAL10 and the XYL1 gene encoding an aldose reductase involved in carbon overflow metabolism. Notably, the regulatory network in C. intermedia, governed by Lac9 and Gal1 from the GALLAC cluster, differs significantly from the (ga)lactose regulons in Saccharomyces cerevisiae, Kluyveromyces lactis and Candida albicans. Moreover, although lactose and galactose metabolism are closely linked in C. intermedia, our results also point to important regulatory differences. This study paves the way to a better understanding of lactose and galactose metabolism in C. intermedia and provides new evolutionary insights into yeast metabolic pathways and regulatory networks. In extension, the results will facilitate future development and use of C. intermedia as a cell-factory for conversion of lactose-rich whey into value-added products.

microbiology↗

bioRxiv2022

Yeasts have evolved divergent enzyme strategies to deconstruct and metabolize xylan

Together with bacteria and fungi, yeasts actively take part in the global carbon cycle. Over a hundred yeast species have been shown to grow on the major plant polysaccharide xylan, which requires an arsenal of carbohydrate active enzymes. However, which enzymatic strategies yeasts use to deconstruct xylan and what specific biological roles they play in its conversion remain unclear. In fact, genome analyses reveal that many xylan-metabolizing yeasts lack expected xylanolytic enzymes. Guided by bioinformatics, we have here selected three xylan-metabolizing ascomycetous yeasts for in-depth characterization of growth behavior and xylanolytic enzymes. The savanna soil yeast Blastobotrys mokoenaii displays superior growth on xylan thanks to an efficient secreted glycoside hydrolase family 11 (GH11) xylanase; solving its crystal structure revealed a high similarity to xylanases from filamentous fungi. The termite gut-associated Scheffersomyces lignosus in contrast grows more slowly and its xylanase activity was found to be mainly cell surface-associated. The wood-isolated Wickerhamomyces canadensis surprisingly could not utilize xylan as the sole carbon source without adding xylooligosaccharides, exogenous xylanases or even by co-culturing with B. mokoenaii, suggesting that W. canadensis relies on initial xylan hydrolysis by neighboring cells. Furthermore, our characterization of a novel W. canadensis GH5 subfamily 49 (GH5_49) xylanase represents the first demonstrated activity in this subfamily. Our collective results provide new information on the variable xylanolytic systems evolved by yeasts and their potential roles in natural carbohydrate conversion. ImportanceMicrobes that take part in the degradation of the polysaccharide xylan, the major hemicellulose component in plant biomass, are equipped with specialized enzyme machineries to hydrolyze the polymer into monosaccharides for further metabolism. However, despite being found in virtually every habitat, little is known of how yeasts break down and metabolize xylan and what biological role they may play in its turnover in nature. Here, we have explored the enzymatic xylan deconstruction strategies of three underexplored yeasts from diverse environments: Blastobotrys mokoenaii from soil, Scheffersomyces lignosus from insect guts and Wickerhamomyces canadensis from trees, and show that each species has a distinct behavior regarding xylan conversion. These findings may be of high relevance for future design and development of microbial cell factories and biorefineries utilizing renewable plant biomass.

microbiology↗