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Redox processes acidify and decarboxylate steam-pretreated lignocellulosic biomass and are modulated by LPMO and catalase.

ABSTRACT: Background:The bioconversion of lignocellulosic feedstocks to ethanol is being commercialised, but further process development is required to improve their economic feasibility. Efficient saccharification of lignocellulose to fermentable sugars requires oxidative cleavage of glycosidic linkages by lytic polysaccharide monooxygenases (LPMOs). However, a proper understanding of the catalytic mechanism of this enzyme class and the interaction with other redox processes associated with the saccharification of lignocellulose is still lacking. The in-use stability of LPMO-containing enzyme cocktails is increased by the addition of catalase implying that hydrogen peroxide (H2O2) is generated in the slurry during incubation. Therefore, we sought to characterize the effects of enzymatic and abiotic sources of H2O2 on lignocellulose hydrolysis to identify parameters that could improve this process. Moreover, we studied the abiotic redox reactions of steam-pretreated wheat straw as a function of temperature and dry-matter (DM) content. Results:Abiotic reactions in pretreated wheat straw consume oxygen, release carbon dioxide (CO2) to the slurry, and decrease the pH. The magnitude of these reactions increased with temperature and with DM content. The presence of LPMO during saccharification reduced the amount of CO2 liberated, while the effect on pH was insignificant. Catalase led to increased decarboxylation through an unknown mechanism. Both in situ-generated and added H2O2 caused a decrease in pH. Conclusions:Abiotic redox processes similar to those that occur in natural water-logged environments also affect the saccharification of pretreated lignocellulose. Heating of the lignocellulosic material and adjustment of pH trigger rapid oxygen consumption and acidification of the slurry. In industrial settings, it will be of utmost importance to control these processes. LPMOs interact with the surrounding redox compounds and redirect abiotic electron flow from decarboxylating reactions to fuel the oxidative cleavage of glycosidic bonds in cellulose.

SUBMITTER: Peciulyte A

PROVIDER: S-EPMC6004669 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Redox processes acidify and decarboxylate steam-pretreated lignocellulosic biomass and are modulated by LPMO and catalase.

Peciulyte Ausra A Samuelsson Louise L Olsson Lisbeth L McFarland K C KC Frickmann Jesper J Østergård Lars L Halvorsen Rune R Scott Brian R BR Johansen Katja S KS

Biotechnology for biofuels 20180618

<h4>Background</h4>The bioconversion of lignocellulosic feedstocks to ethanol is being commercialised, but further process development is required to improve their economic feasibility. Efficient saccharification of lignocellulose to fermentable sugars requires oxidative cleavage of glycosidic linkages by lytic polysaccharide monooxygenases (LPMOs). However, a proper understanding of the catalytic mechanism of this enzyme class and the interaction with other redox processes associated with the s ...[more]

PMID: 29946356

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Project description:BACKGROUND:Non-productive binding of enzymes to lignin is thought to impede the saccharification efficiency of pretreated lignocellulosic biomass to fermentable sugars. Due to a lack of suitable analytical techniques that track binding of individual enzymes within complex protein mixtures and the difficulty in distinguishing the contribution of productive (binding to specific glycans) versus non-productive (binding to lignin) binding of cellulases to lignocellulose, there is currently a poor understanding of individual enzyme adsorption to lignin during the time course of pretreated biomass saccharification. RESULTS:In this study, we have utilized an FPLC (fast protein liquid chromatography)-based methodology to quantify free Trichoderma reesei cellulases (namely CBH I, CBH II, and EG I) concentration within a complex hydrolyzate mixture during the varying time course of biomass saccharification. Three pretreated corn stover (CS) samples were included in this study: Ammonia Fiber Expansion(a) (AFEX™-CS), dilute acid (DA-CS), and ionic liquid (IL-CS) pretreatments. The relative fraction of bound individual cellulases varied depending not only on the pretreated biomass type (and lignin abundance) but also on the type of cellulase. Acid pretreated biomass had the highest levels of non-recoverable cellulases, while ionic liquid pretreated biomass had the highest overall cellulase recovery. CBH II has the lowest thermal stability among the three T. reesei cellulases tested. By preparing recombinant family 1 carbohydrate binding module (CBM) fusion proteins, we have shown that family 1 CBMs are highly implicated in the non-productive binding of full-length T. reesei cellulases to lignin. CONCLUSIONS:Our findings aid in further understanding the complex mechanisms of non-productive binding of cellulases to pretreated lignocellulosic biomass. Developing optimized pretreatment processes with reduced or modified lignin content to minimize non-productive enzyme binding or engineering pretreatment-specific, low-lignin binding cellulases will improve enzyme specific activity, facilitate enzyme recycling, and thereby permit production of cheaper biofuels.

Project description:BackgroundBiogas production from lignocellulosic biomass is generally considered to be challenging due to the recalcitrant nature of this biomass. In this study, the recalcitrance of birch was reduced by applying steam-explosion (SE) pretreatment (210 °C and 10 min). Moreover, bioaugmentation with the cellulolytic bacterium Caldicellulosiruptor bescii was applied to possibly enhance the methane production from steam-exploded birch in an anaerobic digestion (AD) process under thermophilic conditions (62 °C).ResultsOverall, the combined SE and bioaugmentation enhanced the methane yield up to 140% compared to untreated birch, while SE alone contributed to the major share of methane enhancement by 118%. The best methane improvement of 140% on day 50 was observed in bottles fed with pretreated birch and bioaugmentation with lower dosages of C. bescii (2 and 5% of inoculum volume). The maximum methane production rate also increased from 4-mL CH4/g VS (volatile solids)/day for untreated birch to 9-14-mL CH4/g VS/day for steam-exploded birch with applied bioaugmentation. Bioaugmentation was particularly effective for increasing the initial methane production rate of the pretreated birch yielding 21-44% more methane than the pretreated birch without applied bioaugmentation. The extent of solubilization of the organic matter was increased by more than twofold when combined SE pretreatment and bioaugmentation was used in comparison with the methane production from untreated birch. The beneficial effects of SE and bioaugmentation on methane yield indicated that biomass recalcitrance and hydrolysis step are the limiting factors for efficient AD of lignocellulosic biomass. Microbial community analysis by 16S rRNA amplicon sequencing showed that the microbial community composition was altered by the pretreatment and bioaugmentation processes. Notably, the enhanced methane production by pretreatment and bioaugmentation was well correlated with the increase in abundance of key bacterial and archaeal communities, particularly the hydrolytic bacterium Caldicoprobacter, several members of syntrophic acetate oxidizing bacteria and the hydrogenotrophic Methanothermobacter.ConclusionOur findings demonstrate the potential of combined SE and bioaugmentation for enhancing methane production from lignocellulosic biomass.

Project description:BackgroundLignocellulosic biomass is an attractive and sustainable alternative to petroleum-based feedstock for the production of a range of biochemicals, and pretreatment is generally regarded as indispensable for its biorefinery. However, various inhibitors that severely hinder the growth and fermentation of microorganisms are inevitably produced during the pretreatment of lignocellulose. Presently, there are few reports on a single microorganism that can detoxify or tolerate toxic mixtures of pretreated lignocellulose hydrolysate while effectively transforming sugar components into valuable compounds. Alternatively, microbial coculture provides a simpler and more efficacious way to realize this goal by distributing metabolic functions among different specialized strains.ResultsIn this study, a novel synthetic microbial consortium, which is composed of a responsible for detoxification bacterium engineered Pseudomonas putida KT2440 and a lactic acid production specialist Bacillus coagulans NL01, was developed to directly produce lactic acid from highly toxic lignocellulosic hydrolysate. The engineered P. putida with deletion of the sugar metabolism pathway was unable to consume the major fermentable sugars of lignocellulosic hydrolysate but exhibited great tolerance to 10 g/L sodium acetate, 5 g/L levulinic acid, 10 mM furfural and HMF as well as 2 g/L monophenol compound. In addition, the engineered strain rapidly removed diverse inhibitors of real hydrolysate. The degradation rate of organic acids (acetate, levulinic acid) and the conversion rate of furan aldehyde were both 100%, and the removal rate of most monoaromatic compounds remained at approximately 90%. With detoxification using engineered P. putida for 24 h, the 30% (v/v) hydrolysate was fermented to 35.8 g/L lactic acid by B. coagulans with a lactic acid yield of 0.8 g/g total sugars. Compared with that of the single culture of B. coagulans without lactic acid production, the fermentation performance of microbial coculture was significantly improved.ConclusionsThe microbial coculture system constructed in this study demonstrated the strong potential of the process for the biosynthesis of valuable products from lignocellulosic hydrolysates containing high concentrations of complex inhibitors by specifically recruiting consortia of robust microorganisms with desirable characteristics and also provided a feasible and attractive method for the bioconversion of lignocellulosic biomass to other value-added biochemicals.

Dataset Information

Redox processes acidify and decarboxylate steam-pretreated lignocellulosic biomass and are modulated by LPMO and catalase.

Publications

Redox processes acidify and decarboxylate steam-pretreated lignocellulosic biomass and are modulated by LPMO and catalase.

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