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...

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Autores principales:	Bosmat Levi Hevroni, Sarah Moraïs, Yonit Ben-David, Ely Morag, Edward A. Bayer
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	glycoside hydrolases enzymatic profiling scaffoldin cohesin-dockerin specificity CBM cohesin specificity Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/d4836650da17475bbfc404eeb88ddd8e
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spelling	oai:doaj.org-article:d4836650da17475bbfc404eeb88ddd8e2021-11-15T15:57:03ZMinimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content>10.1128/mBio.00443-202150-7511https://doaj.org/article/d4836650da17475bbfc404eeb88ddd8e2020-04-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00443-20https://doaj.org/toc/2150-7511ABSTRACT 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.Bosmat Levi HevroniSarah MoraïsYonit Ben-DavidEly MoragEdward A. BayerAmerican Society for Microbiologyarticleglycoside hydrolasesenzymatic profilingscaffoldincohesin-dockerin specificityCBMcohesin specificityMicrobiologyQR1-502ENmBio, Vol 11, Iss 2 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	glycoside hydrolases enzymatic profiling scaffoldin cohesin-dockerin specificity CBM cohesin specificity Microbiology QR1-502
spellingShingle	glycoside hydrolases enzymatic profiling scaffoldin cohesin-dockerin specificity CBM cohesin specificity Microbiology QR1-502 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>
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.
format	article
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
title	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_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_sort	minimalistic cellulosome of the butanologenic bacterium <named-content content-type="genus-species">clostridium saccharoperbutylacetonicum</named-content>
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/d4836650da17475bbfc404eeb88ddd8e
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Minimalistic Cellulosome of the Butanologenic Bacterium <named-content content-type="genus-species">Clostridium saccharoperbutylacetonicum</named-content>

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