Dataset Information

Isotope-Assisted Metabolite Analysis Sheds Light on Central Carbon Metabolism of a Model Cellulolytic Bacterium Clostridium thermocellum.

ABSTRACT: Cellulolytic bacteria have the potential to perform lignocellulose hydrolysis and fermentation simultaneously. The metabolic pathways of these bacteria, therefore, require more comprehensive and quantitative understanding. Using isotope tracer, gas chromatography-mass spectrometry, and metabolic flux modeling, we decipher the metabolic network of Clostridium thermocellum, a model cellulolytic bacterium which represents as an attractive platform for conversion of lignocellulose to dedicated products. We uncover that the Embden-Meyerhof-Parnas (EMP) pathway is the predominant glycolytic route whereas the Entner-Doudoroff (ED) pathway and oxidative pentose phosphate pathway are inactive. We also observe that C. thermocellum's TCA cycle is initiated by both Si- and Re-citrate synthase, and it is disconnected between 2-oxoglutarate and oxaloacetate in the oxidative direction; C. thermocellum uses a citramalate shunt to synthesize isoleucine; and both the one-carbon pathway and the malate shunt are highly active in this bacterium. To gain a quantitative understanding, we further formulate a fluxome map to quantify the metabolic fluxes through central metabolic pathways. This work represents the first global in vivo investigation of the principal carbon metabolism of C. thermocellum. Our results elucidate the unique structure of metabolic network in this cellulolytic bacterium and demonstrate the capability of isotope-assisted metabolite studies in understanding microbial metabolism of industrial interests.

SUBMITTER: Xiong W

PROVIDER: S-EPMC6115520 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Publications

Isotope-Assisted Metabolite Analysis Sheds Light on Central Carbon Metabolism of a Model Cellulolytic Bacterium <i>Clostridium thermocellum</i>.

Xiong Wei W Lo Jonathan J Chou Katherine J KJ Wu Chao C Magnusson Lauren L Dong Tao T Maness PinChing P

Frontiers in microbiology 20180823

Cellulolytic bacteria have the potential to perform lignocellulose hydrolysis and fermentation simultaneously. The metabolic pathways of these bacteria, therefore, require more comprehensive and quantitative understanding. Using isotope tracer, gas chromatography-mass spectrometry, and metabolic flux modeling, we decipher the metabolic network of <i>Clostridium thermocellum</i>, a model cellulolytic bacterium which represents as an attractive platform for conversion of lignocellulose to dedicate ...[more]

PMID: 30190711

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Project description:BackgroundMicrobial cellulose conversion by Clostridium thermocellum 27405 occurs predominantly through the activity of substrate-adherent bacteria organized in thin, primarily single cell-layered biofilms. The importance of cellulosic surface exposure to microbial hydrolysis has received little attention despite its implied impact on conversion kinetics.ResultsWe showed the spatial heterogeneity of fiber distribution in pure cellulosic sheets, which made direct measurements of biofilm colonization and surface penetration impossible. Therefore, we utilized on-line measurements of carbon dioxide (CO2) production in continuous-flow reactors, in conjunction with confocal imaging, to observe patterns of biofilm invasion and to indirectly estimate microbial accessibility to the substrate's surface and the resulting limitations on conversion kinetics. A strong positive correlation was found between cellulose consumption and CO2 production (R2?=?0.996) and between surface area and maximum biofilm activity (R2?=?0.981). We observed an initial biofilm development rate (0.46 h-1, 0.34 h-1 and 0.33 h-1) on Whatman sheets (#1, #598 and #3, respectively) that stabilized when the accessible surface was maximally colonized. The results suggest that cellulose conversion kinetics is initially subject to a microbial limitation period where the substrate is in excess, followed by a substrate limitation period where cellular mass, in the form of biofilms, is not limiting. Accessible surface area acts as an important determinant of the respective lengths of these two distinct periods. At end-point fermentation, all sheets were digested predominantly under substrate accessibility limitations (e.g., up to 81% of total CO2 production for Whatman #1). Integration of CO2 production rates over time showed Whatman #3 underwent the fastest conversion efficiency under microbial limitation, suggestive of best biofilm penetration, while Whatman #1 exhibited the least recalcitrance and the faster degradation during the substrate limitation period.ConclusionThe results showed that the specific biofilm development rate of cellulolytic bacteria such as C. thermocellum has a notable effect on overall reactor kinetics during the period of microbial limitation, when ca. 20% of cellulose conversion occurs. The study further demonstrated the utility of on-line CO2 measurements as a method to assess biofilm development and substrate digestibility pertaining to microbial solubilization of cellulose, which is relevant when considering feedstock pre-treatment options.

Dataset Information

Isotope-Assisted Metabolite Analysis Sheds Light on Central Carbon Metabolism of a Model Cellulolytic Bacterium Clostridium thermocellum.

Publications

Isotope-Assisted Metabolite Analysis Sheds Light on Central Carbon Metabolism of a Model Cellulolytic Bacterium <i>Clostridium thermocellum</i>.

Similar Datasets

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