Elucidating Xylose Metabolism of Scheffersomyces stipitis for Lignocellulosic Ethanol Production

Abstract

The conversion of pentose to ethanol is one of the major barriers of industrializing the lignocellulosic ethanol processes. As one of the most promising native strains for pentose fermentation, Scheffersomyces stipitis (formerly known as Pichia stipitis) has been widely studied for its xylose fermentation. In spite of the abundant experimental evidence regarding ethanol and byproducts production under various aeration conditions, the mathematical descriptions of the processes are rare. In this work, a constraint-based metabolic network model for the central carbon metabolism of S. stipitis was reconstructed by integrating genomic (S. stipitis v2.0, KEGG), biochemical (ChEBI, PubChem), and physiological information available for this microorganism and other related yeast. The model consists of the stoichiometry of metabolic reactions, biosynthetic requirements for growth, and other constraints. Flux balance analysis is applied to characterize the phenotypic behavior of S. stipitis grown on xylose. The model predictions are in good agreement with published experimental results. To understand the effect of redox balance on xylose fermentation, we propose a system identification-based metabolic analysis framework to extract biological knowledge embedded in a series of designed in silico experiments. In the proposed framework, we first design in silico experiments to perturb the metabolic network in order to investigate the interested properties and then perform system identification, whereby applying principal component analysis (PCA) to the data generated by the designed in silico experiments. By combining the in silico perturbation experiments with system identification tools, biologically meaningful information contained in the complex network structure can be decomposed and translated into easily interpretable information that is useful for biologist. The PCA analysis identifies the phenotypic changes caused by oxygen supply and reveals key metabolic reactions related to redox homeostasis in different phenotypes. In addition, the influence of the cofactor preference of key enzyme (xylose reductase) in xylose metabolism is investigated using the proposed approach, and the results provide important insights on cofactor engineering of xylose metabolism.

Promiscuous activities of heterologous enzymes lead to unintended metabolic rerouting in Saccharomyces cerevisiae engineered to assimilate various sugars from renewable biomass

“…1 c) was probably due to the significant accumulation of galactitol and d -tagatose. For assimilation of xylose, there exist two major pathways: the oxidoreductive pathway composed of XR and XDH, mainly found in fungi such as S. stipitis [ 8 ] and the isomerase pathway composed of xylose isomerase, mainly found in bacteria such as Clostridium phytofermentans [ 24 ] and anaerobic fungi [ 25 ]. In particular, XR (EC.…”

“…In our previous studies, S. cerevisiae has been successfully engineered to catabolize xylose and cellobiose simultaneously (resulting in strain EJ4) using multiple genetic perturbations and rational and laboratory evolution, as follows [ 7 ]. Initially, the oxidoreductive xylose-assimilating pathway consisting of XYL1 , XYL2 , and XYL3 coding for xylose reductase (XR), xylitol dehydrogenase (XDH), and xylulokinase, respectively, from Scheffersomyces stipitis [ 8 ] was introduced into S. cerevisiae D452-2 by multicopy integration (resulting in strain SR7) [ 9 ]. Then, strain SR8, a fast and efficient xylose-fermenting strain, was derived from strain SR7 through serial subculturing of strain SR7 in the presence of xylose followed by deletion of ALD6 coding for acetaldehyde dehydrogenase to minimize acetate production [ 9 ].…”

Effects of aeration on growth, ethanol and polyol accumulation by Spathaspora passalidarum NRRL Y‐27907 and Scheffersomyces stipitis NRRL Y‐7124

“…During xylose assimilation, S. stipitis ' xylose reductase (XR) can use either NADH or NADPH as a cofactor, but it has a higher affinity for NADPH. The next enzyme in this pathway, xylitol dehydrogenase (XDH), which converts xylitol into xylulose, depends on NAD + almost exclusively (Liang et al, ). The different reactivities of the two enzymes with their respective cofactors can deplete NAD + and accumulate NADH.…”

Fermentation Process and Metabolic Flux of Ethanol Production from the Detoxified Hydrolyzate of Cassava Residue

“…Furthermore, the effects of dissolved oxygen on the fermentation of yeast have been studied. Appropriate amount of dissolved oxygen is found to be an important control parameter for ethanol fermentation; in particular, it strongly influences redox balance, cell growth, and energy generation for xylose transport (Liang et al, 2014 ).…”