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Exploring Metabolism in the Extremel...

Rubinstein, Gabriel Michael.

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Exploring Metabolism in the Extremely Thermophilic Cellulolytic Bacterium Caldicellulosiruptor Bescii for Engineered Bioproduct Formation.

紀錄類型:	書目-電子資源 : Monograph/item
正題名/作者:	Exploring Metabolism in the Extremely Thermophilic Cellulolytic Bacterium Caldicellulosiruptor Bescii for Engineered Bioproduct Formation./
作者:	Rubinstein, Gabriel Michael.
出版者:	Ann Arbor : ProQuest Dissertations & Theses, : 2020,
面頁冊數:	305 p.
附註:	Source: Dissertations Abstracts International, Volume: 82-08, Section: B.
Contained By:	Dissertations Abstracts International82-08B.
標題:	Biochemistry. -
電子資源:	https://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=28154652
ISBN:	9798569917297

Exploring Metabolism in the Extremely Thermophilic Cellulolytic Bacterium Caldicellulosiruptor Bescii for Engineered Bioproduct Formation.
Rubinstein, Gabriel Michael.

Exploring Metabolism in the Extremely Thermophilic Cellulolytic Bacterium Caldicellulosiruptor Bescii for Engineered Bioproduct Formation. - Ann Arbor : ProQuest Dissertations & Theses, 2020 - 305 p.

Source: Dissertations Abstracts International, Volume: 82-08, Section: B.

Thesis (Ph.D.)--University of Georgia, 2020.

This item must not be sold to any third party vendors.

Petroleum is the primary source of liquid transportation fuels and industrial chemicals, such as solvents and plastic precursors. Due to the finite nature of oil and the need to mitigate environmental damage from the combustion of hydrocarbons, renewable methods for producing these chemicals are necessary. One approach is to generate liquid fuels and commodity chemicals from the microbial conversion of lignocellulosic biomass, an abundant and renewable feedstock. Before lignocellulose can be utilized by most microorganisms, extensive pretreatment is necessary to release the monosaccharides for fermentation. The use of biomass degrading organisms for biological conversion can decrease costs associated with pretreatment. Caldicellulosiruptor bescii is an extremely thermophilic anaerobic Gram-positive bacterium with the highest optimum growth temperature of any known cellulolytic organism (78 °C). Due to the availability of genetic tools and the unusual capacity to grow on completely untreated lignocellulosic biomass as a sole carbon substrate, C. bescii has gained attention as a potential platform organism to produce biofuels and other bioproducts via consolidated bioprocessing. The work described herein examines metabolism in wild type C. bescii, strains containing targeted gene deletions, and strains expressing engineered metabolic pathways. Study of native metabolism revealed the presence of an alternative enzyme for the oxidation of glyceraldehyde-3-phosphate in glycolysis. This enzyme, a new family of tungsten-containing glyceraldehyde-3-phosphate oxidoreductase, was purified and a knockout strain was characterized. Bioinformatic analysis of the genome revealed a multipurpose ATPase (msmK) involved in sugar uptake. A deletion strain showed that msmK helps transport di- and oligosaccharides but not C5 and C6 substrates. A two-gene engineered metabolic pathway for the reduction of carboxylic acids to alcohols was expressed in C. bescii, the first instance of this pathway in a cellulolytic organism. Consisting of an aldehyde oxidoreductase and a primary alcohol dehydrogenase, it enabled the reduction of exogenously added organic acids to the corresponding alcohol, with concomitant ethanol generation from acetate. Finally, heterologous expression of a pyruvate oxidoreductase resulted in increased pyruvate-oxidizing activity in cytosolic extracts. Together, this work describes advances in the understanding of redox cofactor balancing, carbohydrate utilization, carbon metabolism, and heterologous gene expression in this biotechnologically-relevant bacterium.

ISBN: 9798569917297Subjects--Topical Terms:

518028
Biochemistry.
Subjects--Index Terms:

Bacteria

Exploring Metabolism in the Extremely Thermophilic Cellulolytic Bacterium Caldicellulosiruptor Bescii for Engineered Bioproduct Formation.
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520 $a Petroleum is the primary source of liquid transportation fuels and industrial chemicals, such as solvents and plastic precursors. Due to the finite nature of oil and the need to mitigate environmental damage from the combustion of hydrocarbons, renewable methods for producing these chemicals are necessary. One approach is to generate liquid fuels and commodity chemicals from the microbial conversion of lignocellulosic biomass, an abundant and renewable feedstock. Before lignocellulose can be utilized by most microorganisms, extensive pretreatment is necessary to release the monosaccharides for fermentation. The use of biomass degrading organisms for biological conversion can decrease costs associated with pretreatment. Caldicellulosiruptor bescii is an extremely thermophilic anaerobic Gram-positive bacterium with the highest optimum growth temperature of any known cellulolytic organism (78 °C). Due to the availability of genetic tools and the unusual capacity to grow on completely untreated lignocellulosic biomass as a sole carbon substrate, C. bescii has gained attention as a potential platform organism to produce biofuels and other bioproducts via consolidated bioprocessing. The work described herein examines metabolism in wild type C. bescii, strains containing targeted gene deletions, and strains expressing engineered metabolic pathways. Study of native metabolism revealed the presence of an alternative enzyme for the oxidation of glyceraldehyde-3-phosphate in glycolysis. This enzyme, a new family of tungsten-containing glyceraldehyde-3-phosphate oxidoreductase, was purified and a knockout strain was characterized. Bioinformatic analysis of the genome revealed a multipurpose ATPase (msmK) involved in sugar uptake. A deletion strain showed that msmK helps transport di- and oligosaccharides but not C5 and C6 substrates. A two-gene engineered metabolic pathway for the reduction of carboxylic acids to alcohols was expressed in C. bescii, the first instance of this pathway in a cellulolytic organism. Consisting of an aldehyde oxidoreductase and a primary alcohol dehydrogenase, it enabled the reduction of exogenously added organic acids to the corresponding alcohol, with concomitant ethanol generation from acetate. Finally, heterologous expression of a pyruvate oxidoreductase resulted in increased pyruvate-oxidizing activity in cytosolic extracts. Together, this work describes advances in the understanding of redox cofactor balancing, carbohydrate utilization, carbon metabolism, and heterologous gene expression in this biotechnologically-relevant bacterium.
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