Metagenomics for the development of new biocatalysts to advance lignocellulose saccharification for bioeconomic development

Abstract

The world economy is moving toward the use of renewable and nonedible lignocellulosic biomasses as substitutes for fossil sources in order to decrease the environmental impact of manufacturing processes and overcome the conflict with food production. Enzymatic hydrolysis of the feedstock is a key technology for bio-based chemical production, and the identification of novel, less expensive and more efficient biocatalysts is one of the main challenges. As the genomic era has shown that only a few microorganisms can be cultured under standard laboratory conditions, the extraction and analysis of genetic material directly from environmental samples, termed metagenomics, is a promising way to overcome this bottleneck. Two screening methodologies can be used on metagenomic material: the function-driven approach of expression libraries and sequence-driven analysis based on gene homology. Both techniques have been shown to be useful for the discovery of novel biocatalysts for lignocellulose conversion, and they enabled identification of several (hemi)cellulases and accessory enzymes involved in (hemi)cellulose hydrolysis. This review summarizes the latest progress in metagenomics aimed at discovering new enzymes for lignocellulose saccharification.

Applying functional metagenomics to search for novel lignocellulosic enzymes in a microbial consortium derived from a thermophilic composting phase of sugarcane bagasse and cow manure

“…The identification of new hydrolytic enzymes or genetic improvement of existing ones could help lower the costs of enzyme production for biomass degradation, and there is considerable effort being directed toward improving cellulosic ethanol production by these and other means (Himmel et al 2007;Duan et al 2009;Li et al 2009;Gnansounou and Dauriat 2010;Horn et al 2012;Montella et al 2015).…”

A Rapid Method for the Selection of Amidohydrolases from Metagenomic Libraries by Applying Synthetic Nucleosides and a Uridine Auxotrophic Host

“…The hydrolytic activity of the recombinant enzymes was assayed qualitatively by testing 29 different substrates ( Figure 4A): N 4 -acylated (2'-deoxy)cytidines and cytosines (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16); N 2 -acetylisocytosines (17); chromogenic substrates such as p-nitroacetanilide (18) and p-nitrobenzanilide (19); various p-nitrophenyl (pNP) esters (20)(21)(22)(23)(24); terephthalate derivatives bis(2-hydroxyethyl) terephthalate (26), dimethyl terephthalate (27), and monomethyl terephthalate (28); and the beta-lactam nitrocefin (29). Hydrolysis of chromogenic substrates (18)(19)(20)(21)(22)(23)(24)(25) was analysed spectrophotometrically at 405 nm, and that of other substrates was analysed spectrophotometrically at 240-320 nm and using TLC or HPLC-MS ( Figures S4-S10).…”

“…Keeping in mind that the functions of 30%-40% of the genes in genomes remain unknown, the detection of new biocatalysts in the sequenced genomes can be difficult if no significant homology with known enzymes is observed [6,7]. Metagenomics, the improvement of high-throughput screening methods, and the development of host expression systems for metagenome-derived genes, systems biology, and gene synthesis may together open the gateway to the useful information that hides in unexplored genetic resources, e.g., new enzymes with unique properties and novel scaffolds applicable for evolution in vitro [8][9][10][11][12].…”

From lignocellulosic metagenomes to lignocellulolytic genes: trends, challenges and future prospects

“…Metagenomic screening of a switch grass adapted community for thermotolerant enzymes yielded 21 suitable candidates, of which those with a T opt >70% correlated with a 78% chance of being co‐tolerant to high strength ionic liquids . Further metagenomics‐driven investigations of thermophilic systems are likely to progress this area, particularly when coupled with directed evolution strategies for enzyme optimisation as reviewed recently . The findings highlight the importance of careful sampling of target environments, such as, for example sugarcane bagasse, not only from a substrate perspective re lignocellulosic material abundance, but also to take account of variations in micro‐environmental conditions .…”