Expression of a hyperthermophilic endoglucanase in hybrid poplar modifies the plant cell wall and enhances digestibility

Abstract

BackgroundExpression of glycosyl hydrolases in lignocellulosic biomass has been proposed as an alternative to improve efficiency of cellulosic ethanol production. In planta production of hyperthermophilic hydrolytic enzymes could prevent the detrimental effects often seen resulting from the expression of recombinant mesophilic enzymes to plant hosts. Utilizing lignocellulosic feedstocks to produce hyperthermophilic hydrolases provides additional benefits for ethanol production in the way of transgenic feedstocks serving as both enzyme providers and cellulosic substrates.ResultsIn this study, transgenic hybrid poplar (Populus alba × grandidentata) was generated to express a hyperthermophilic endoglucanase from Thermotoga neapolitana with an optimal temperature over 100 °C. Functional hyperthermoactive endoglucanase was successfully produced in the transgenic events, and altered phenotypic growth was observed in transgenic lines. Moreover, the line with the highest TnCelB expression in both leaf and developing xylem had reduced lignin content and cellulose crystallinity, resulting in a more digestible cell wall. The activation of TnCelB by a post-harvest heat treatment resulted in enhanced saccharification efficiencies of transgenic poplar lines with moderate TnCelB expression and without alteration of cellulose and lignin when not heat-treated. In planta high-level overexpression of a hyperthermophilic endoglucanase paired with heat treatment following harvest, resulted in biomass that was comparable with wild-type lines that underwent a traditional pretreatment for saccharification.ConclusionsOverexpression of hyperthermophilic endoglucanase in feedstock had impacts on plant growth and cell wall composition, especially when the enzyme was highly expressed. Improved glucan saccharification efficiencies from transgenic lines before and after heat treatment could reduce both the economic and environmental costs associated with ethanol production from lignocellulosic biomass.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1224-7) contains supplementary material, which is available to authorized users.

Engineering hydroxyproline‐O‐glycosylated biopolymers to reconstruct the plant cell wall for improved biomass processability

“…Reduced recalcitrance of plant biomass by in planta expression of E1cd and other CWD enzymes has been shown in many previous reports (Brunecky et al, 2011; Chou et al, 2011; Chung et al, 2015; Donohoe et al, 2017; Klose et al, 2015; Mir et al, 2017; Xiao et al, 2018). It has been demonstrated that the decreased recalcitrance was not due to the postpretreatment residual enzyme activity.…”

“…Alternatively, key cell wall‐depolymerizing (CWD) enzymes, such as a cellulase or xylanase, have been introduced into plants to improve postharvest biomass processability (Brunecky et al, 2011; Chou, Dai, Hsieh, & Ku, 2011; Donohoe et al, 2017; Klose, Roder, Girfoglio, Fischer, & Commandeur, 2012). This would allow the biomass feedstocks to serve as both enzyme suppliers and substrates for ethanol production (Xiao, He, Ojeda‐Lassalle, Poovaiah, & Coleman, 2018). To avoid detrimental effects on the growth of plant hosts, thermophilic or hyperthermophilic enzymes are often used for expression in planta.…”

Targeted genome editing of plants and plant cells for biomanufacturing

“…Plants can be more suitable for the production of exocellulases than microorganisms due to the higher enzyme activity (Klinger et al 2015). Endoglucanases can be secreted to the apoplast (Xiao et al 2018), but targeting to the ER (Klose et al 2015) or plastids (Schmidt et al 2019b;Fumagalli et al 2019) helps to prevent the enzymes degrading essential cellulose structures before plants are harvested and homogenized for product extraction. Alternatively, enzyme expression can be restricted to specific organs, such as maize kernels (Vicuna Requesens et al 2019), or can be induced at a particular time, such as just before harvest, thus reducing the fiber content and rigidity, which can facilitate primary product extraction.…”

Xylan in the Middle: Understanding Xylan Biosynthesis and Its Metabolic Dependencies Toward Improving Wood Fiber for Industrial Processing

“…Such transgenic approaches have not only produced favorable phenotypes for industrial processing, but have resulted in plants with novel SCW properties as well as additional traits such as increased stress tolerance. Self-processing plants with increased industrial processing efficiency and biotic defense responses are generated by in planta expression of xylan targeting CWDE from wood rotting fungi that can be active under normal plant growth (Gandla et al, 2014; Pawar et al, 2017b), or only become active upon application of external stimulus (Shen et al, 2012; Mir et al, 2017; Xiao et al, 2018; Table 1). Various studies have shown that xylan modifications differ between plant species and lineages (Allerdings et al, 2006; Rennie and Scheller, 2014; Peña et al, 2016; Smith et al, 2017; de Souza et al, 2018) and modification genes from one lineage can add modifications to the xylan backbone of another (Anders et al, 2012; Xiong et al, 2015; Zhong et al, 2018; Table 1).…”