Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome

Abstract Background Carbohydrate-active enzymes (CAZymes) form the most widespread and structurally diverse set of enzymes involved in the breakdown, biosynthesis, or modification of lignocellulose that can be found in living organisms. However, the structural diversity of CAZymes has rendered the t...

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Autores principales:	André L. A. Neves, Jiangkun Yu, Yutaka Suzuki, Marisol Baez-Magana, Elena Arutyunova, Eóin O’Hara, Tim McAllister, Kim H. Ominski, M. Joanne Lemieux, Le Luo Guan
Formato:	article
Lenguaje:	EN
Publicado:	BMC 2021
Materias:	Cattle Feed efficiency Microbial enzymes Rumen microbiota Microbial ecology QR100-130
Acceso en línea:	https://doaj.org/article/c16a0943fa1049f685c4114b526e2347
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spelling	oai:doaj.org-article:c16a0943fa1049f685c4114b526e23472021-11-28T12:08:14ZAccelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome10.1186/s40168-021-01147-12049-2618https://doaj.org/article/c16a0943fa1049f685c4114b526e23472021-11-01T00:00:00Zhttps://doi.org/10.1186/s40168-021-01147-1https://doaj.org/toc/2049-2618Abstract Background Carbohydrate-active enzymes (CAZymes) form the most widespread and structurally diverse set of enzymes involved in the breakdown, biosynthesis, or modification of lignocellulose that can be found in living organisms. However, the structural diversity of CAZymes has rendered the targeted discovery of novel enzymes extremely challenging, as these proteins catalyze many different chemical reactions and are sourced by a vast array of microbes. Consequently, many uncharacterized members of CAZyme families of interest have been overlooked by current methodologies (e.g., metagenomic screening) used to discover lignocellulolytic enzymes. Results In the present study, we combined phenotype-based selective pressure on the rumen microbiota with targeted functional profiling to guide the discovery of unknown CAZymes. In this study, we found 61 families of glycoside hydrolases (GH) (out of 182 CAZymes) from protein sequences deposited in the CAZy database—currently associated with more than 20,324 microbial genomes. Phenotype-based selective pressure on the rumen microbiome showed that lignocellulolytic bacteria (e.g., Fibrobacter succinogenes, Butyrivibrio proteoclasticus) and three GH families (e.g., GH11, GH13, GH45) exhibited an increased relative abundance in the rumen of feed efficient cattle when compared to their inefficient counterparts. These results paved the way for the application of targeted functional profiling to screen members of the GH11 and GH45 families against a de novo protein reference database comprised of 1184 uncharacterized enzymes, which led to the identification of 18 putative xylanases (GH11) and three putative endoglucanases (GH45). The biochemical proof of the xylanolytic activity of the newly discovered enzyme validated the computational simulations and demonstrated the stability of the most abundant xylanase. Conclusions These findings contribute to the discovery of novel enzymes for the breakdown, biosynthesis, or modification of lignocellulose and demonstrate that the rumen microbiome is a source of promising enzyme candidates for the biotechnology industry. The combined approaches conceptualized in this study can be adapted to any microbial environment, provided that the targeted microbiome is easy to manipulate and facilitates enrichment for the microbes of interest. Video AbstractAndré L. A. NevesJiangkun YuYutaka SuzukiMarisol Baez-MaganaElena ArutyunovaEóin O’HaraTim McAllisterKim H. OminskiM. Joanne LemieuxLe Luo GuanBMCarticleCattleFeed efficiencyMicrobial enzymesRumen microbiotaMicrobial ecologyQR100-130ENMicrobiome, Vol 9, Iss 1, Pp 1-16 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Cattle Feed efficiency Microbial enzymes Rumen microbiota Microbial ecology QR100-130
spellingShingle	Cattle Feed efficiency Microbial enzymes Rumen microbiota Microbial ecology QR100-130 André L. A. Neves Jiangkun Yu Yutaka Suzuki Marisol Baez-Magana Elena Arutyunova Eóin O’Hara Tim McAllister Kim H. Ominski M. Joanne Lemieux Le Luo Guan Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
description	Abstract Background Carbohydrate-active enzymes (CAZymes) form the most widespread and structurally diverse set of enzymes involved in the breakdown, biosynthesis, or modification of lignocellulose that can be found in living organisms. However, the structural diversity of CAZymes has rendered the targeted discovery of novel enzymes extremely challenging, as these proteins catalyze many different chemical reactions and are sourced by a vast array of microbes. Consequently, many uncharacterized members of CAZyme families of interest have been overlooked by current methodologies (e.g., metagenomic screening) used to discover lignocellulolytic enzymes. Results In the present study, we combined phenotype-based selective pressure on the rumen microbiota with targeted functional profiling to guide the discovery of unknown CAZymes. In this study, we found 61 families of glycoside hydrolases (GH) (out of 182 CAZymes) from protein sequences deposited in the CAZy database—currently associated with more than 20,324 microbial genomes. Phenotype-based selective pressure on the rumen microbiome showed that lignocellulolytic bacteria (e.g., Fibrobacter succinogenes, Butyrivibrio proteoclasticus) and three GH families (e.g., GH11, GH13, GH45) exhibited an increased relative abundance in the rumen of feed efficient cattle when compared to their inefficient counterparts. These results paved the way for the application of targeted functional profiling to screen members of the GH11 and GH45 families against a de novo protein reference database comprised of 1184 uncharacterized enzymes, which led to the identification of 18 putative xylanases (GH11) and three putative endoglucanases (GH45). The biochemical proof of the xylanolytic activity of the newly discovered enzyme validated the computational simulations and demonstrated the stability of the most abundant xylanase. Conclusions These findings contribute to the discovery of novel enzymes for the breakdown, biosynthesis, or modification of lignocellulose and demonstrate that the rumen microbiome is a source of promising enzyme candidates for the biotechnology industry. The combined approaches conceptualized in this study can be adapted to any microbial environment, provided that the targeted microbiome is easy to manipulate and facilitates enrichment for the microbes of interest. Video Abstract
format	article
author	André L. A. Neves Jiangkun Yu Yutaka Suzuki Marisol Baez-Magana Elena Arutyunova Eóin O’Hara Tim McAllister Kim H. Ominski M. Joanne Lemieux Le Luo Guan
author_facet	André L. A. Neves Jiangkun Yu Yutaka Suzuki Marisol Baez-Magana Elena Arutyunova Eóin O’Hara Tim McAllister Kim H. Ominski M. Joanne Lemieux Le Luo Guan
author_sort	André L. A. Neves
title	Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
title_short	Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
title_full	Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
title_fullStr	Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
title_full_unstemmed	Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
title_sort	accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome
publisher	BMC
publishDate	2021
url	https://doaj.org/article/c16a0943fa1049f685c4114b526e2347
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Accelerated discovery of novel glycoside hydrolases using targeted functional profiling and selective pressure on the rumen microbiome

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