Expanded genome and proteome reallocation in a novel, robust Bacillus coagulans strain capable of utilizing pentose and hexose sugars

David Dooley; Seunghyun Ryu; Richard J. Giannone; Jackson Edwards; Bruce S. Dien; Patricia J. Slininger; Cong T. Trinh

doi:10.1128/msystems.00952-24

Expanded genome and proteome reallocation in a novel, robust Bacillus coagulans strain capable of utilizing pentose and hexose sugars

David Dooley, Seunghyun Ryu, Richard J. Giannone, Jackson Edwards, Bruce S. Dien, Patricia J. Slininger, Cong T. Trinh

Bioanalytical Mass Spectrometry

Research output: Contribution to journal › Article › peer-review

Abstract

Bacillus coagulans, a Gram-positive thermophilic bacterium, is recognized for its probiotic properties and recent development as a microbial cell factory. Despite its importance for biotechnological applications, the current understanding of B. coagulans’ robustness is limited, especially for undomesticated strains. To fill this knowledge gap, we characterized the metabolic capability and performed functional genomics and systems analysis of a novel, robust strain, B. coagulans B-768. Genome sequencing revealed that B-768 has the largest B. coagulans genome known to date (3.94 Mbp), about 0.63 Mbp larger than the average genome of sequenced B. coagulans strains, with expanded carbohydrate metabolism and mobilome. Functional genomics identified a well-equipped genetic portfolio for utilizing a wide range of C5 (xylose, arabinose), C6 (glucose, mannose, galactose), and C12 (cellobiose) sugars present in biomass hydrolysates, which was validated experimentally. For growth on individual xylose and glucose, the dominant sugars in biomass hydrolysates, B-768 exhibited distinct phenotypes and proteome profiles. Faster growth and glucose uptake rates resulted in lactate overflow metabolism, which makes B. coagulans a lactate overproducer; however, slower growth and xylose uptake diminished overflow metabolism due to the high energy demand for sugar assimilation. Carbohydrate Transport and Metabolism (COG-G), Translation (COG-J), and Energy Conversion and Production (COG-C) made up 60%–65% of the measured proteomes but were allocated differently when growing on xylose and glucose. The trade-off in proteome reallocation, with high investment in COG-C over COG-G, explains the xylose growth phenotype with significant upregulation of xylose metabolism, pyruvate metabolism, and tricarboxylic acid (TCA) cycle. Strain B-768 tolerates and effectively utilizes inhibitory biomass hydrolysates containing mixed sugars and exhibits hierarchical sugar utilization with glucose as the preferential substrate.

Original language	English
Journal	mSystems
Volume	9
Issue number	11
DOIs	https://doi.org/10.1128/msystems.00952-24
State	Published - Nov 2024

Funding

This research was funded by the DOE BER award (DE-SC0022226 to C.T.T., R.J.G., and B.S.D.). The sequencing project was supported by the Center for Bioenergy Innovation (CBI), which is a U.S. Department of Energy Bioenergy Research Center supported by the of Biological and Environmental Research in the DOE of Science. Oak Ridge National Laboratory is managed by UT-Battelle, LLC for the US DOE under Contract Number DE-AC05-00OR22725. The work (Project IDs # 510751, 509417) conducted by the U.S. Department of Energy Joint Genome Institute, a DOE of Science User Facility, is supported under Contract No. DE-AC02-05CH11231. B.S.D., J.E., and P.J.S. received support from the U.S. Department of Agriculture, Agricultural Research Service (CRIS Numbers 5010-41000-189). This research was funded by the DOE BER award (DE-SC0022226 to C.T.T., R.J.G., and B.S.D.). The sequencing project was supported by the Center for Bioenergy Innovation (CBI), which is a U.S. Department of Energy Bioenergy Research Center supported by the Office of Biological and Environmental Research in the DOE Office of Science. Oak Ridge National Laboratory is managed by UT-Battelle, LLC for the US DOE under Contract Number DE-AC05-00OR22725. The work (Project IDs # 510751, 509417) conducted by the U.S. Department of Energy Joint Genome Institute, a DOE Office of Science User Facility, is supported under Contract No. DE-AC02-05CH11231. B.S.D., J.E., and P.J.S. received financial support from the U.S. Department of Agriculture, Agricultural Research Service (CRIS Numbers 5010-41000-189). The mention of trade names or commercial products in this article is solely to provide scientific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. USDA is an equal opportunity provider and employer.

Funders	Funder number
Agricultural Research Service
U.S. Department of Energy Bioenergy Research Center
Biological and Environmental Research
Oak Ridge National Laboratory
Center for Bioenergy Innovation
DOE of Science
U.S. Department of Agriculture
U.S. Department of Energy Joint Genome Institute	DE-AC02-05CH11231
Crown Research Institutes	5010-41000-189
U.S. Department of Energy	DE-AC05-00OR22725, 509417, 510751, DE-SC0022226
Office of Science	DE-AC02-05CH11231

Keywords

B-768
Bacillus coagulans
biomass hydrolysates
carbohydrate metabolism
lactate overproducer
overflow metabolism
proteome reallocation
robustness
switchgrass
thermophile

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1128/msystems.00952-24

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title = "Expanded genome and proteome reallocation in a novel, robust Bacillus coagulans strain capable of utilizing pentose and hexose sugars",

abstract = "Bacillus coagulans, a Gram-positive thermophilic bacterium, is recognized for its probiotic properties and recent development as a microbial cell factory. Despite its importance for biotechnological applications, the current understanding of B. coagulans{\textquoteright} robustness is limited, especially for undomesticated strains. To fill this knowledge gap, we characterized the metabolic capability and performed functional genomics and systems analysis of a novel, robust strain, B. coagulans B-768. Genome sequencing revealed that B-768 has the largest B. coagulans genome known to date (3.94 Mbp), about 0.63 Mbp larger than the average genome of sequenced B. coagulans strains, with expanded carbohydrate metabolism and mobilome. Functional genomics identified a well-equipped genetic portfolio for utilizing a wide range of C5 (xylose, arabinose), C6 (glucose, mannose, galactose), and C12 (cellobiose) sugars present in biomass hydrolysates, which was validated experimentally. For growth on individual xylose and glucose, the dominant sugars in biomass hydrolysates, B-768 exhibited distinct phenotypes and proteome profiles. Faster growth and glucose uptake rates resulted in lactate overflow metabolism, which makes B. coagulans a lactate overproducer; however, slower growth and xylose uptake diminished overflow metabolism due to the high energy demand for sugar assimilation. Carbohydrate Transport and Metabolism (COG-G), Translation (COG-J), and Energy Conversion and Production (COG-C) made up 60%–65% of the measured proteomes but were allocated differently when growing on xylose and glucose. The trade-off in proteome reallocation, with high investment in COG-C over COG-G, explains the xylose growth phenotype with significant upregulation of xylose metabolism, pyruvate metabolism, and tricarboxylic acid (TCA) cycle. Strain B-768 tolerates and effectively utilizes inhibitory biomass hydrolysates containing mixed sugars and exhibits hierarchical sugar utilization with glucose as the preferential substrate.",

keywords = "B-768, Bacillus coagulans, biomass hydrolysates, carbohydrate metabolism, lactate overproducer, overflow metabolism, proteome reallocation, robustness, switchgrass, thermophile",

author = "David Dooley and Seunghyun Ryu and Giannone, {Richard J.} and Jackson Edwards and Dien, {Bruce S.} and Slininger, {Patricia J.} and Trinh, {Cong T.}",

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AU - Dooley, David

AU - Ryu, Seunghyun

AU - Giannone, Richard J.

AU - Edwards, Jackson

AU - Dien, Bruce S.

AU - Slininger, Patricia J.

AU - Trinh, Cong T.

PY - 2024/11

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N2 - Bacillus coagulans, a Gram-positive thermophilic bacterium, is recognized for its probiotic properties and recent development as a microbial cell factory. Despite its importance for biotechnological applications, the current understanding of B. coagulans’ robustness is limited, especially for undomesticated strains. To fill this knowledge gap, we characterized the metabolic capability and performed functional genomics and systems analysis of a novel, robust strain, B. coagulans B-768. Genome sequencing revealed that B-768 has the largest B. coagulans genome known to date (3.94 Mbp), about 0.63 Mbp larger than the average genome of sequenced B. coagulans strains, with expanded carbohydrate metabolism and mobilome. Functional genomics identified a well-equipped genetic portfolio for utilizing a wide range of C5 (xylose, arabinose), C6 (glucose, mannose, galactose), and C12 (cellobiose) sugars present in biomass hydrolysates, which was validated experimentally. For growth on individual xylose and glucose, the dominant sugars in biomass hydrolysates, B-768 exhibited distinct phenotypes and proteome profiles. Faster growth and glucose uptake rates resulted in lactate overflow metabolism, which makes B. coagulans a lactate overproducer; however, slower growth and xylose uptake diminished overflow metabolism due to the high energy demand for sugar assimilation. Carbohydrate Transport and Metabolism (COG-G), Translation (COG-J), and Energy Conversion and Production (COG-C) made up 60%–65% of the measured proteomes but were allocated differently when growing on xylose and glucose. The trade-off in proteome reallocation, with high investment in COG-C over COG-G, explains the xylose growth phenotype with significant upregulation of xylose metabolism, pyruvate metabolism, and tricarboxylic acid (TCA) cycle. Strain B-768 tolerates and effectively utilizes inhibitory biomass hydrolysates containing mixed sugars and exhibits hierarchical sugar utilization with glucose as the preferential substrate.

AB - Bacillus coagulans, a Gram-positive thermophilic bacterium, is recognized for its probiotic properties and recent development as a microbial cell factory. Despite its importance for biotechnological applications, the current understanding of B. coagulans’ robustness is limited, especially for undomesticated strains. To fill this knowledge gap, we characterized the metabolic capability and performed functional genomics and systems analysis of a novel, robust strain, B. coagulans B-768. Genome sequencing revealed that B-768 has the largest B. coagulans genome known to date (3.94 Mbp), about 0.63 Mbp larger than the average genome of sequenced B. coagulans strains, with expanded carbohydrate metabolism and mobilome. Functional genomics identified a well-equipped genetic portfolio for utilizing a wide range of C5 (xylose, arabinose), C6 (glucose, mannose, galactose), and C12 (cellobiose) sugars present in biomass hydrolysates, which was validated experimentally. For growth on individual xylose and glucose, the dominant sugars in biomass hydrolysates, B-768 exhibited distinct phenotypes and proteome profiles. Faster growth and glucose uptake rates resulted in lactate overflow metabolism, which makes B. coagulans a lactate overproducer; however, slower growth and xylose uptake diminished overflow metabolism due to the high energy demand for sugar assimilation. Carbohydrate Transport and Metabolism (COG-G), Translation (COG-J), and Energy Conversion and Production (COG-C) made up 60%–65% of the measured proteomes but were allocated differently when growing on xylose and glucose. The trade-off in proteome reallocation, with high investment in COG-C over COG-G, explains the xylose growth phenotype with significant upregulation of xylose metabolism, pyruvate metabolism, and tricarboxylic acid (TCA) cycle. Strain B-768 tolerates and effectively utilizes inhibitory biomass hydrolysates containing mixed sugars and exhibits hierarchical sugar utilization with glucose as the preferential substrate.

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Expanded genome and proteome reallocation in a novel, robust Bacillus coagulans strain capable of utilizing pentose and hexose sugars

Abstract

Funding

Keywords

UN SDGs

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