Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature

ABSTRACT Designer cellulosomes consist of chimeric cohesin-bearing scaffoldins for the controlled incorporation of recombinant dockerin-containing enzymes. The largest designer cellulosome reported to date is a chimeric scaffoldin that contains 6 cohesins. This scaffoldin represented a technical lim...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Johanna Stern, Sarah Moraïs, Raphael Lamed, Edward A. Bayer
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/329e87803b994ee8a7706595dc747b03
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:329e87803b994ee8a7706595dc747b03
record_format	dspace
spelling	oai:doaj.org-article:329e87803b994ee8a7706595dc747b032021-11-15T15:41:41ZAdaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature10.1128/mBio.00083-162150-7511https://doaj.org/article/329e87803b994ee8a7706595dc747b032016-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00083-16https://doaj.org/toc/2150-7511ABSTRACT Designer cellulosomes consist of chimeric cohesin-bearing scaffoldins for the controlled incorporation of recombinant dockerin-containing enzymes. The largest designer cellulosome reported to date is a chimeric scaffoldin that contains 6 cohesins. This scaffoldin represented a technical limit of sorts, since adding another cohesin proved problematic, owing to resultant low expression levels, instability (cleavage) of the scaffoldin polypeptide, and limited numbers of available cohesin-dockerin specificities—the hallmark of designer cellulosomes. Nevertheless, increasing the number of enzymes integrated into designer cellulosomes is critical, in order to further enhance degradation of plant cell wall material. Adaptor scaffoldins comprise an intermediate type of scaffoldin that can both incorporate various enzymes and attach to an additional scaffoldin. Using this strategy, we constructed an efficient form of adaptor scaffoldin that possesses three type I cohesins for enzyme integration, a single type II dockerin for interaction with an additional scaffoldin, and a carbohydrate-binding module for targeting to the cellulosic substrate. In parallel, we designed a hexavalent scaffoldin capable of connecting to the adaptor scaffoldin by the incorporation of an appropriate type II cohesin. The resultant extended designer cellulosome comprised 8 recombinant enzymes—4 xylanases and 4 cellulases—thereby representing a potent enzymatic cocktail for solubilization of natural lignocellulosic substrates. The contribution of the adaptor scaffoldin clearly demonstrated that proximity between the two scaffoldins and their composite set of enzymes is crucial for optimized degradation. After 72 h of incubation, the performance of the extended designer cellulosome was determined to be approximately 70% compared to that of native cellulosomes. IMPORTANCE Plant cell wall residues represent a major source of renewable biomass for the production of biofuels such as ethanol via breakdown to soluble sugars. The natural microbial degradation process, however, is inefficient for achieving cost-effective processes in the conversion of plant-derived biomass to biofuels, either from dedicated crops or human-generated cellulosic wastes. The accumulation of the latter is considered a major environmental pollutant. The development of designer cellulosome nanodevices for enhanced plant cell wall degradation thus has major impacts in the fields of environmental pollution, bioenergy production, and biotechnology in general. The findings reported in this article comprise a true breakthrough in our capacity to produce extended designer cellulosomes via synthetic biology means, thus enabling the assembly of higher-order complexes that can supersede the number of enzymes included in a single multienzyme complex.Johanna SternSarah MoraïsRaphael LamedEdward A. BayerAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 7, Iss 2 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Johanna Stern Sarah Moraïs Raphael Lamed Edward A. Bayer Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
description	ABSTRACT Designer cellulosomes consist of chimeric cohesin-bearing scaffoldins for the controlled incorporation of recombinant dockerin-containing enzymes. The largest designer cellulosome reported to date is a chimeric scaffoldin that contains 6 cohesins. This scaffoldin represented a technical limit of sorts, since adding another cohesin proved problematic, owing to resultant low expression levels, instability (cleavage) of the scaffoldin polypeptide, and limited numbers of available cohesin-dockerin specificities—the hallmark of designer cellulosomes. Nevertheless, increasing the number of enzymes integrated into designer cellulosomes is critical, in order to further enhance degradation of plant cell wall material. Adaptor scaffoldins comprise an intermediate type of scaffoldin that can both incorporate various enzymes and attach to an additional scaffoldin. Using this strategy, we constructed an efficient form of adaptor scaffoldin that possesses three type I cohesins for enzyme integration, a single type II dockerin for interaction with an additional scaffoldin, and a carbohydrate-binding module for targeting to the cellulosic substrate. In parallel, we designed a hexavalent scaffoldin capable of connecting to the adaptor scaffoldin by the incorporation of an appropriate type II cohesin. The resultant extended designer cellulosome comprised 8 recombinant enzymes—4 xylanases and 4 cellulases—thereby representing a potent enzymatic cocktail for solubilization of natural lignocellulosic substrates. The contribution of the adaptor scaffoldin clearly demonstrated that proximity between the two scaffoldins and their composite set of enzymes is crucial for optimized degradation. After 72 h of incubation, the performance of the extended designer cellulosome was determined to be approximately 70% compared to that of native cellulosomes. IMPORTANCE Plant cell wall residues represent a major source of renewable biomass for the production of biofuels such as ethanol via breakdown to soluble sugars. The natural microbial degradation process, however, is inefficient for achieving cost-effective processes in the conversion of plant-derived biomass to biofuels, either from dedicated crops or human-generated cellulosic wastes. The accumulation of the latter is considered a major environmental pollutant. The development of designer cellulosome nanodevices for enhanced plant cell wall degradation thus has major impacts in the fields of environmental pollution, bioenergy production, and biotechnology in general. The findings reported in this article comprise a true breakthrough in our capacity to produce extended designer cellulosomes via synthetic biology means, thus enabling the assembly of higher-order complexes that can supersede the number of enzymes included in a single multienzyme complex.
format	article
author	Johanna Stern Sarah Moraïs Raphael Lamed Edward A. Bayer
author_facet	Johanna Stern Sarah Moraïs Raphael Lamed Edward A. Bayer
author_sort	Johanna Stern
title	Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
title_short	Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
title_full	Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
title_fullStr	Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
title_full_unstemmed	Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature
title_sort	adaptor scaffoldins: an original strategy for extended designer cellulosomes, inspired from nature
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/329e87803b994ee8a7706595dc747b03
work_keys_str_mv	AT johannastern adaptorscaffoldinsanoriginalstrategyforextendeddesignercellulosomesinspiredfromnature AT sarahmorais adaptorscaffoldinsanoriginalstrategyforextendeddesignercellulosomesinspiredfromnature AT raphaellamed adaptorscaffoldinsanoriginalstrategyforextendeddesignercellulosomesinspiredfromnature AT edwardabayer adaptorscaffoldinsanoriginalstrategyforextendeddesignercellulosomesinspiredfromnature
_version_	1718427634768019456

Adaptor Scaffoldins: An Original Strategy for Extended Designer Cellulosomes, Inspired from Nature

Ejemplares similares