Dataset Information

Enhanced delignification of steam-pretreated poplar by a bacterial laccase.

ABSTRACT: The recalcitrance of woody biomass, particularly its lignin component, hinders its sustainable transformation to fuels and biomaterials. Although the recent discovery of several bacterial ligninases promises the development of novel biocatalysts, these enzymes have largely been characterized using model substrates: direct evidence for their action on biomass is lacking. Herein, we report the delignification of woody biomass by a small laccase (sLac) from Amycolatopsis sp. 75iv3. Incubation of steam-pretreated poplar (SPP) with sLac enhanced the release of acid-precipitable polymeric lignin (APPL) by ~6-fold, and reduced the amount of acid-soluble lignin by ~15%. NMR spectrometry revealed that the APPL was significantly syringyl-enriched relative to the original material (~16:1 vs. ~3:1), and that sLac preferentially oxidized syringyl units and altered interunit linkage distributions. sLac's substrate preference among monoaryls was also consistent with this observation. In addition, sLac treatment reduced the molar mass of the APPL by over 50%, as determined by gel-permeation chromatography coupled with multi-angle light scattering. Finally, sLac acted synergistically with a commercial cellulase cocktail to increase glucose production from SPP ~8%. Overall, this study establishes the lignolytic activity of sLac on woody biomass and highlights the biocatalytic potential of bacterial enzymes.

SUBMITTER: Singh R

PROVIDER: S-EPMC5294454 | biostudies-other | 2017 Feb

REPOSITORIES: biostudies-other

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Enhanced delignification of steam-pretreated poplar by a bacterial laccase.

Singh Rahul R Hu Jinguang J Regner Matthew R MR Round James W JW Ralph John J Saddler John N JN Eltis Lindsay D LD

Scientific reports 20170207

The recalcitrance of woody biomass, particularly its lignin component, hinders its sustainable transformation to fuels and biomaterials. Although the recent discovery of several bacterial ligninases promises the development of novel biocatalysts, these enzymes have largely been characterized using model substrates: direct evidence for their action on biomass is lacking. Herein, we report the delignification of woody biomass by a small laccase (sLac) from Amycolatopsis sp. 75iv3. Incubation of st ...[more]

PMID: 28169340

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Project description: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.

Project description:Background:The current production of bioethanol based on lignocellulosic biomass (LCB) highly depends on thermostable enzymes and extremophiles owing to less risk of contamination. Thermophilic bacterial cellulases are preferred over fungi due to their higher growth rate, presence of complex multi-enzymes, stability, and enhanced bioconversion efficiency. Corncob, underutilized biomass, ensures energy conservation due to high lignocellulosic and more fermentable sugar content. In the present study, the thermophilic bacterium Bacillus aerius CMCPS1, isolated from the thermal springs of Manikaran, Himachal Pradesh, India, was characterized in terms of its activity, stability, and hydrolytic capacity. A two-step process comprising: (i) a combined strategy of hydrodynamic cavitation reaction (HCR)-coupled enzymatic (LccH at 6.5 U) pretreatment for delignification and (ii) subsequent hydrolysis of pre-treated (HCR-LccH) corncob biomass (CCB) using a thermostable cocktail of CMCPS1 was adopted to validate the efficiency of the process. Some of the parameters studied include lignin reduction, cellulose increase, and saccharification efficiency. Result:Among the five isolates obtained by in situ enrichment on various substrates, B. aerius CMCPS1, isolated from hot springs, exhibited the maximum hydrolytic activity of 4.11. The GH activity of the CMCPS1 strain under submerged fermentation revealed maximum filter paper activity (FPA) and endoglucanase activity of 4.36 IU mL-1 and 2.98 IU mL-1, respectively, at 44 h. Similarly, the isolate produced exoglucanase and ?-glucosidase with an activity of 1.76 IU mL-1 and 1.23 IU mL-1 at 48 h, respectively. More specifically, the enzyme endo-1,4-?-d glucanase E.C.3.2.1.4 (CMCase) produced by B. aerius CMCPS1 displayed wider stability to pH (3-9) and temperature (30-90 °C) than most fungal cellulases. Similarly, the activity of CMCase increased in the presence of organic solvents (118% at 30% acetone v/v). The partially purified CMCase from the culture supernatant of CMCPS1 registered 64% yield with twofold purification. The zymogram and SDS-PAGE analyses further confirmed the CMCase activity with an apparent molecular mass of 70 kDa. The presence of genes specific to cellulases, such as cellulose-binding domain CelB, confirmed the presence of GH family 46 and ?-glucosidase activity (GH3). The multifunctional cellulases of CMCPS1 were evaluated for their saccharification efficiency on laccase (LccH, a fungal laccase from Hexagonia hirta MSF2)-pretreated corncob in a HCR. The lignin and hemicelluloses removal efficiency of HCR-LccH was 54.1 and 6.57%, respectively, with an increase in cellulose fraction (42.25%). The saccharification efficiency of 55% was achieved with CMCPS1 multifunctional cellulases at 50 °C and pH 5.0. Conclusion:The multifunctional cellulase complex of B. aerius CMCPS1 is a potential biocatalyst for application in lignocellulosic biomass-based biorefineries. The saccharification ability of HCR-LccH-pretreated corncob at elevated temperatures would be an advantage for biofuel production from lignocellulosic biomass.

Dataset Information

Enhanced delignification of steam-pretreated poplar by a bacterial laccase.

Publications

Enhanced delignification of steam-pretreated poplar by a bacterial laccase.

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