Profiling glucose-induced selective inhibition of disaccharide catabolism in Bacillus megaterium QM B1551 by stable isotope labelling

Youngster, Tracy; Wushensky, Julie A.; Aristilde, Ludmilla

doi:10.1099/mic.0.000540

Cited by 4 publications

(7 citation statements)

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“…Much of what is known about carbon metabolism in Bacillus species is derived from Bacillus subtilis , the primary model organism for Gram-positive bacteria ( Sauer et al, 1997 ; Dauner and Sauer, 2001 ; Dauner et al, 2001 ; Fuhrer et al, 2005 ). Previous metabolic studies highlighted incongruent metabolic fluxes between B. subtilis and B. megaterium species ( Sauer et al, 1997 ; Dauner and Sauer, 2001 ; Dauner et al, 2001 ; Fuhrer et al, 2005 ; Furch et al, 2007a , b ; Tannler et al, 2008 ; Youngster et al, 2017 ). Metabolic flux modeling has been performed on mutant strains of B. megaterium that lacked specific metabolic functions ( Furch et al, 2007b ).…”

Section: Introductionmentioning

confidence: 99%

“…Glucose, a monomer of the biopolymers cellulose and starch, is a common carbohydrate in environmental matrices ( Koegel-Knabner, 2002 ). The disaccharide sucrose, a dimer with glucose and fructose, is common in plant materials and can serve as a carbon source to B. megaterium ( Youngster et al, 2017 ). Xylose, a pentose monosaccharide, is a major monomer in hemicellulose, an abundant component of plant cell walls ( Koegel-Knabner, 2002 ).…”

Section: Introductionmentioning

confidence: 99%

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Flux Connections Between Gluconate Pathway, Glycolysis, and Pentose–Phosphate Pathway During Carbohydrate Metabolism in Bacillus megaterium QM B1551

et al. 2018

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Bacillus megaterium is a bacterium of great importance as a plant-beneficial bacterium in agricultural applications and in industrial bioproduction of proteins. Understanding intracellular processing of carbohydrates in this species is crucial to predicting natural carbon utilization as well as informing strategies in metabolic engineering. Here, we applied stable isotope-assisted metabolomics profiling and metabolic flux analysis to elucidate, at high resolution, the connections of the different catabolic routes for carbohydrate metabolism immediately following substrate uptake in B. megaterium QM B1551. We performed multiple 13C tracer experiments to obtain both kinetic and long-term 13C profiling of intracellular metabolites. In addition to the direct phosphorylation of glucose to glucose-6-phosphate (G6P) prior to oxidation to 6-phosphogluconate (6P-gluconate), the labeling data also captured glucose catabolism through the gluconate pathway involving glucose oxidation to gluconate followed by phosphorylation to 6P-gluconate. Our data further confirmed the absence of the Entner–Doudoroff pathway in B. megaterium and showed that subsequent catabolism of 6P-gluconate was instead through the oxidative pentose–phosphate (PP) pathway. Quantitative flux analysis of glucose-grown cells showed equal partition of consumed glucose from G6P to the Embden–Meyerhof–Parnas (EMP) pathway and from G6P to the PP pathway through 6P-gluconate. Growth on fructose alone or xylose alone was consistent with the ability of B. megaterium to use each substrate as a sole source of carbon. However, a detailed 13C mapping during simultaneous feeding of B. megaterium on glucose, fructose, and xylose indicated non-uniform intracellular investment of the different carbohydrate substrates. Flux of glucose-derived carbons dominated the gluconate pathway and the PP pathway, whereas carbon flux from both glucose and fructose populated the EMP pathway; there was no assimilatory flux of xylose-derived carbons. Collectively, our findings provide new quantitative insights on the contribution of the different catabolic routes involved in initiating carbohydrate catabolism in B. megaterium and related Bacillus species.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Flux Connections Between Gluconate Pathway, Glycolysis, and Pentose–Phosphate Pathway During Carbohydrate Metabolism in Bacillus megaterium QM B1551

et al. 2018

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“…The trace metal concentrations were as follows: 0.96 μM CuSO 4 ·5H 2 O, 0.97 μM H 3 BO 3 , 9.5 μM ZnSO 4 ·7H 2 O, 0.87 μM MnSO 4 ·5H 2 O, 0.21 μM NiCl 2 ·6H 2 O, and 0.34 μM Na 2 MoO 4 ·5H 2 O. Succinate (200 mM C) was provided as the carbon source for P. protegens Pf-5, and glucose (55 mM C) was provided as the carbon source for P. megaterium QM B1551.…”

Section: Methodsmentioning

confidence: 99%

“…Cell cultures (three biological replicates) were grown at 30 °C in a Thermo Scientific SHKE6000-7 incubator shaker (Thermo Fisher Scientific Waltham, MA) at 220 rpm. 53,54 The growth medium, which was pH-adjusted (pH 7.0) and filter-sterilized (0.22 μm nylon; MilliporeSigma, Darmstadt, Germany), contained the following major salts: 20 53 was provided as the carbon source for P. protegens Pf-5, and glucose (55 mM C) 54 was provided as the carbon source for P. megaterium QM B1551. The cells were grown under iron (Fe)-replete conditions with 30 μM total dissolved unchelated Fe or under Fedeficient conditions with 50 nM total dissolved unchelated Fe.…”

Section: Culturing Conditionsmentioning

confidence: 99%

Effects of Phosphonate Herbicides on the Secretions of Plant-Beneficial Compounds by Two Plant Growth-Promoting Soil Bacteria: A Metabolomics Investigation

Wilkes

Aristilde

2021

ACS Environ. Au

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Plant growth-promoting rhizobacteria (PGPR) that colonize plant roots produce a variety of plant-beneficial compounds, including plant-growth regulators, metal-scavenging compounds, and antibiotics against plant pathogens. Adverse effects of phosphonate herbicides, the most extensively used herbicides, on the growth and metabolism of PGPR species have been widely reported. However, the potential consequence of these effects on the biosynthesis and secretion of PGPR-derived beneficial compounds still remains to be investigated. Here, using high-resolution mass spectrometry and a metabolomics approach, we investigated both the intracellular metabolome and the extracellular secretions of biomass-normalized metabolite levels in two PGPR species (Pseudomonas protegens Pf-5, a Gram-negative bacterium; Priestia megaterium QM B1551, a Grampositive bacterium) exposed to three common phosphonate herbicides (glyphosate, glufosinate, and fosamine; 0.1−1 mM) in either iron (Fe)-replete or Fe-deficient nutrient media. We quantified secreted auxin-type plant hormone compounds (phenylacetic acid and indole-3-acetic acid), iron-scavenging compounds or siderophores (pyoverdine and schizokinen), and antibiotics (2,4diacetylphloroglucinol and pyoluteorin) produced by these PGPR species. The Fe-replete cells exposed to the phosphonate herbicides yielded up to a 25-fold increase in the production of both auxin and antibiotic compounds, indicating that herbicide exposure under Fe-replete conditions triggered metabolite secretions. However, the herbicide-exposed Fe-deficient cells exhibited a near 2-fold depletion in the secretion of these auxin and antibiotic compounds as well as a 77% decrease in siderophore production. Intracellular metabolomics analysis of the Fe-deficient cells further revealed metabolic perturbations in biosynthetic pathways consistent with the impaired production of the plant-beneficial compounds. Our findings implied that compromised cellular metabolism during nutrient deficiency may exacerbate the adverse effects of phosphonate herbicides on PGPR species.

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“…P. megaterium can be found in diverse habits including honey (López and Alippi 2009 ), wine (von Cosmos et al 2017 ), raw meat (Yucel et al 2009 ), fish (Al Bulushi et al 2010 ), sea water (Xu et al 2014 ), the oral cavity of humans (Al-Thubiani et al 2018 ), and most typically plants and soil (Dobrzanski et al 2018 ). Consequently, its metabolism is adapted to a variety of different carbon sources including xylose (a byproduct of hemicellulose), glycerol (de Jesus et al 2016 ; Korneli et al 2013 ; Moreno et al 2015 ), disaccharides such as cellobiose, maltose or sucrose (Youngster et al 2017 ), and a range of cheap mixed saccharide sources such as sugarcane molasses (Kanjanachumpol et al 2013 ).…”

Section: Priestia Megaterium —History and Genome Sequencingmentioning

confidence: 99%

The “beauty in the beast”—the multiple uses of Priestia megaterium in biotechnology

Biedendieck

Knuuti

Jahn

2021

Appl Microbiol Biotechnol

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Over 30 years, the Gram-positive bacterium Priestia megaterium (previously known as Bacillus megaterium) was systematically developed for biotechnological applications ranging from the production of small molecules like vitamin B12, over polymers like polyhydroxybutyrate (PHB) up to the in vivo and in vitro synthesis of multiple proteins and finally whole-cell applications. Here we describe the use of the natural vitamin B12 (cobalamin) producer P. megaterium for the elucidation of the biosynthetic pathway and the subsequent systematic knowledge-based development for production purposes. The formation of PHB, a natural product of P. megaterium and potential petro-plastic substitute, is covered and discussed. Further important biotechnological characteristics of P. megaterium for recombinant protein production including high protein secretion capacity and simple cultivation on value-added carbon sources are outlined. This includes the advanced system with almost 30 commercially available expression vectors for the intracellular and extracellular production of recombinant proteins at the g/L scale. We also revealed a novel P. megaterium transcription-translation system as a complementary and versatile biotechnological tool kit. As an impressive biotechnology application, the formation of various cytochrome P450 is also critically highlighted. Finally, whole cellular applications in plant protection are completing the overall picture of P. megaterium as a versatile giant cell factory. Key points • The use of Priestia megaterium for the biosynthesis of small molecules and recombinant proteins through to whole-cell applications is reviewed. • P. megaterium can act as a promising alternative host in biotechnological production processes.

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Profiling glucose-induced selective inhibition of disaccharide catabolism in Bacillus megaterium QM B1551 by stable isotope labelling

Cited by 4 publications

References 28 publications

Flux Connections Between Gluconate Pathway, Glycolysis, and Pentose–Phosphate Pathway During Carbohydrate Metabolism in Bacillus megaterium QM B1551

Flux Connections Between Gluconate Pathway, Glycolysis, and Pentose–Phosphate Pathway During Carbohydrate Metabolism in Bacillus megaterium QM B1551

Effects of Phosphonate Herbicides on the Secretions of Plant-Beneficial Compounds by Two Plant Growth-Promoting Soil Bacteria: A Metabolomics Investigation

The “beauty in the beast”—the multiple uses of Priestia megaterium in biotechnology

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