Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content>
ABSTRACT Clostridium saccharoperbutylacetonicum is a mesophilic, anaerobic, butanol-producing bacterium, originally isolated from soil. It was recently reported that C. saccharoperbutylacetonicum possesses multiple cellulosomal elements and would potentially form the smallest cellulosome known in na...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2020-04-01
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| Series: | mBio |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mBio.00443-20 |
| _version_ | 1818434521471647744 |
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| author | Bosmat Levi Hevroni Sarah Moraïs Yonit Ben-David Ely Morag Edward A. Bayer |
| author_facet | Bosmat Levi Hevroni Sarah Moraïs Yonit Ben-David Ely Morag Edward A. Bayer |
| author_sort | Bosmat Levi Hevroni |
| collection | DOAJ |
| description | ABSTRACT Clostridium saccharoperbutylacetonicum is a mesophilic, anaerobic, butanol-producing bacterium, originally isolated from soil. It was recently reported that C. saccharoperbutylacetonicum possesses multiple cellulosomal elements and would potentially form the smallest cellulosome known in nature. Its genome contains only eight dockerin-bearing enzymes, and its unique scaffoldin bears two cohesins (Cohs), three X2 modules, and two carbohydrate-binding modules (CBMs). In this study, all of the cellulosome-related modules were cloned, expressed, and purified. The recombinant cohesins, dockerins, and CBMs were tested for binding activity using enzyme-linked immunosorbent assay (ELISA)-based techniques. All the enzymes were tested for their comparative enzymatic activity on seven different cellulosic and hemicellulosic substrates, thus revealing four cellulases, a xylanase, a mannanase, a xyloglucanase, and a lichenase. All dockerin-containing enzymes interacted similarly with the second cohesin (Coh2) module, whereas Coh1 was more restricted in its interaction pattern. In addition, the polysaccharide-binding properties of the CBMs within the scaffoldin were examined by two complementary assays, affinity electrophoresis and affinity pulldown. The scaffoldin of C. saccharoperbutylacetonicum exhibited high affinity for cellulosic and hemicellulosic substrates, specifically to microcrystalline cellulose and xyloglucan. Evidence that supports substrate-dependent in vivo secretion of cellulosomes is presented. The results of our analyses contribute to a better understanding of simple cellulosome systems by identifying the key players in this minimalistic system and the binding pattern of its cohesin-dockerin interaction. The knowledge gained by our study will assist further exploration of similar minimalistic cellulosomes and will contribute to the significance of specific sets of defined cellulosomal enzymes in the degradation of cellulosic biomass. IMPORTANCE Cellulosome-producing bacteria are considered among the most important bacteria in both mesophilic and thermophilic environments, owing to their capacity to deconstruct recalcitrant plant-derived polysaccharides (and notably cellulose) into soluble saccharides for subsequent processing. In many ecosystems, the cellulosome-producing bacteria are particularly effective “first responders.” The massive amounts of sugars produced are potentially amenable in industrial settings to further fermentation by appropriate microbes to biofuels, notably ethanol and butanol. Among the solvent-producing bacteria, Clostridium saccharoperbutylacetonicum has the smallest cellulosome system known thus far. The importance of investigating the building blocks of such a small, multifunctional nanomachine is crucial to understanding the fundamental activities of this efficient enzymatic complex. |
| first_indexed | 2024-12-14T16:38:19Z |
| format | Article |
| id | doaj.art-d4836650da17475bbfc404eeb88ddd8e |
| institution | Directory Open Access Journal |
| issn | 2150-7511 |
| language | English |
| last_indexed | 2024-12-14T16:38:19Z |
| publishDate | 2020-04-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mBio |
| spelling | doaj.art-d4836650da17475bbfc404eeb88ddd8e2022-12-21T22:54:24ZengAmerican Society for MicrobiologymBio2150-75112020-04-0111210.1128/mBio.00443-20Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content>Bosmat Levi Hevroni0Sarah Moraïs1Yonit Ben-David2Ely Morag3Edward A. Bayer4Department of Biomolecular Sciences, The Weizmann Institute of Science, Rehovot, IsraelDepartment of Biomolecular Sciences, The Weizmann Institute of Science, Rehovot, IsraelDepartment of Biomolecular Sciences, The Weizmann Institute of Science, Rehovot, IsraelDepartment of Biomolecular Sciences, The Weizmann Institute of Science, Rehovot, IsraelDepartment of Biomolecular Sciences, The Weizmann Institute of Science, Rehovot, IsraelABSTRACT Clostridium saccharoperbutylacetonicum is a mesophilic, anaerobic, butanol-producing bacterium, originally isolated from soil. It was recently reported that C. saccharoperbutylacetonicum possesses multiple cellulosomal elements and would potentially form the smallest cellulosome known in nature. Its genome contains only eight dockerin-bearing enzymes, and its unique scaffoldin bears two cohesins (Cohs), three X2 modules, and two carbohydrate-binding modules (CBMs). In this study, all of the cellulosome-related modules were cloned, expressed, and purified. The recombinant cohesins, dockerins, and CBMs were tested for binding activity using enzyme-linked immunosorbent assay (ELISA)-based techniques. All the enzymes were tested for their comparative enzymatic activity on seven different cellulosic and hemicellulosic substrates, thus revealing four cellulases, a xylanase, a mannanase, a xyloglucanase, and a lichenase. All dockerin-containing enzymes interacted similarly with the second cohesin (Coh2) module, whereas Coh1 was more restricted in its interaction pattern. In addition, the polysaccharide-binding properties of the CBMs within the scaffoldin were examined by two complementary assays, affinity electrophoresis and affinity pulldown. The scaffoldin of C. saccharoperbutylacetonicum exhibited high affinity for cellulosic and hemicellulosic substrates, specifically to microcrystalline cellulose and xyloglucan. Evidence that supports substrate-dependent in vivo secretion of cellulosomes is presented. The results of our analyses contribute to a better understanding of simple cellulosome systems by identifying the key players in this minimalistic system and the binding pattern of its cohesin-dockerin interaction. The knowledge gained by our study will assist further exploration of similar minimalistic cellulosomes and will contribute to the significance of specific sets of defined cellulosomal enzymes in the degradation of cellulosic biomass. IMPORTANCE Cellulosome-producing bacteria are considered among the most important bacteria in both mesophilic and thermophilic environments, owing to their capacity to deconstruct recalcitrant plant-derived polysaccharides (and notably cellulose) into soluble saccharides for subsequent processing. In many ecosystems, the cellulosome-producing bacteria are particularly effective “first responders.” The massive amounts of sugars produced are potentially amenable in industrial settings to further fermentation by appropriate microbes to biofuels, notably ethanol and butanol. Among the solvent-producing bacteria, Clostridium saccharoperbutylacetonicum has the smallest cellulosome system known thus far. The importance of investigating the building blocks of such a small, multifunctional nanomachine is crucial to understanding the fundamental activities of this efficient enzymatic complex.https://journals.asm.org/doi/10.1128/mBio.00443-20glycoside hydrolasesenzymatic profilingscaffoldincohesin-dockerin specificityCBMcohesin specificity |
| spellingShingle | Bosmat Levi Hevroni Sarah Moraïs Yonit Ben-David Ely Morag Edward A. Bayer Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> mBio glycoside hydrolases enzymatic profiling scaffoldin cohesin-dockerin specificity CBM cohesin specificity |
| title | Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> |
| title_full | Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> |
| title_fullStr | Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> |
| title_full_unstemmed | Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> |
| title_short | Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content> |
| title_sort | minimalistic cellulosome of the butanologenic bacterium named content content type genus species clostridium saccharoperbutylacetonicum named content |
| topic | glycoside hydrolases enzymatic profiling scaffoldin cohesin-dockerin specificity CBM cohesin specificity |
| url | https://journals.asm.org/doi/10.1128/mBio.00443-20 |
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