scholarly journals Physiological Characteristics and Comparative Secretome Analysis of Morchella importuna Grown on Glucose, Rice Straw, Sawdust, Wheat Grain, and MIX Substrates

2021 ◽  
Vol 12 ◽  
Author(s):  
YingLi Cai ◽  
XiaoLong Ma ◽  
QianQian Zhang ◽  
FuQiang Yu ◽  
Qi Zhao ◽  
...  
Keyword(s):  
Rice Straw ◽  
Lignin Degradation ◽  
Extracellular Enzymes ◽  
Plant Biomass ◽  
Growth Potential ◽  
Glycoside Hydrolases ◽  
Lignocellulose Degradation ◽  
Wheat Grain ◽  
Predominant Component ◽  
Morchella Importuna

Morels (Morchella sp.) are economically important edible macro-fungi, which can grow on various synthetic or semi-synthetic media. However, the complex nutritional metabolism and requirements of these fungi remain ill-defined. This study, based on the plant biomass commonly used in the artificial cultivation of morels, assessed and compared the growth characteristics and extracellular enzymes of Morchella importuna cultivated on glucose, rice straw, sawdust, wheat grain, and a mixture of equal proportions of the three latter plant substrates (MIX). M. importuna could grow on all five tested media but displayed significant variations in mycelial growth rate, biomass, and sclerotium yield on the different media. The most suitable medium for M. importuna was wheat and wheat-containing medium, followed by glucose, while rice straw and sawdust were the least suitable. A total of 268 secretory proteins were identified by liquid chromatography coupled with tandem mass spectrometry detection. Functional classification and label-free comparative analysis of these proteins revealed that carbohydrate-active enzyme (CAZYme) proteins were the predominant component of the secretome of M. importuna, followed by protease, peptidase, and other proteins. The abundances of CAZYme proteins differed among the tested media, ranging from 64% on glucose to 88% on rice straw. The CAZYme classes of glycoside hydrolases and carbohydrate-binding module were enriched in the five secretomes. Furthermore, the enzyme activities of CMCase, lignase, amylase, xylase, pNPCase, and pNPGase were detected during the continuous culture of M. importuna in MIX medium, and the relative expression of the corresponding genes were detected by quantitative real-time PCR. The combined data of growth potential, secretome, extracellular enzyme activity, and gene expression on different substrates inferred that M. importuna was weak in lignocellulose degradation but a good starch decomposer. Specifically, in terms of the degradation of cellulose, the ability to degrade cellulose into oligosaccharides was weaker compared with further degradation into monosaccharides, and this might be the speed-limiting step of cellulose utilization in M. importuna. In addition, M. importuna had a strong ability to decompose various hemicellulose glycosidic bonds, especially α- and β-galactosidase. Only a very few lignin-degradation-related proteins were detected, and these were in low abundance, consistent with the presence of weak lignin degradation ability. Furthermore, the presence of lipase and chitinase implied that M. importuna was capable of decomposition of its own mycelia in vitro. The study provides key data that facilitates a further understanding of the complex nutritional metabolism of M. importuna.

scholarly journals Genus-Wide Assessment of Lignocellulose Utilization in the Extremely Thermophilic GenusCaldicellulosiruptorby Genomic, Pangenomic, and Metagenomic Analyses

2018 ◽  
Vol 84 (9) ◽  
Author(s):  
Laura L. Lee ◽  
Sara E. Blumer-Schuette ◽  
Javier A. Izquierdo ◽  
Jeffrey V. Zurawski ◽  
Andrew J. Loder ◽  
...  
Keyword(s):  
Microcrystalline Cellulose ◽  
Plant Biomass ◽  
Thermophilic Bacteria ◽  
Cellulose Degradation ◽  
Glycoside Hydrolases ◽  
Metagenomic Data ◽  
Lignocellulose Degradation ◽  
Sequence Information ◽  
Metagenomic Sequence ◽  
Content Type

ABSTRACTMetagenomic data from Obsidian Pool (Yellowstone National Park, USA) and 13 genome sequences were used to reassess genus-wide biodiversity for the extremely thermophilicCaldicellulosiruptor. The updated core genome contains 1,401 ortholog groups (average genome size for 13 species = 2,516 genes). The pangenome, which remains open with a revised total of 3,493 ortholog groups, encodes a variety of multidomain glycoside hydrolases (GHs). These include three cellulases with GH48 domains that are colocated in the glucan degradation locus (GDL) and are specific determinants for microcrystalline cellulose utilization. Three recently sequenced species,Caldicellulosiruptorsp. strain Rt8.B8 (renamed hereCaldicellulosiruptor morganii),Thermoanaerobacter cellulolyticusstrain NA10 (renamed hereCaldicellulosiruptor naganoensis), andCaldicellulosiruptorsp. strain Wai35.B1 (renamed hereCaldicellulosiruptor danielii), degraded Avicel and lignocellulose (switchgrass).C. morganiiwas more efficient thanCaldicellulosiruptor besciiin this regard and differed from the other 12 species examined, both based on genome content and organization and in the specific domain features of conserved GHs. Metagenomic analysis of lignocellulose-enriched samples from Obsidian Pool revealed limited new information on genus biodiversity. Enrichments yielded genomic signatures closely related to that ofCaldicellulosiruptor obsidiansis, but there was also evidence for other thermophilic fermentative anaerobes (Caldanaerobacter,Fervidobacterium,Caloramator, andClostridium). One enrichment, containing 89.8%Caldicellulosiruptorand 9.7%Caloramator, had a capacity for switchgrass solubilization comparable to that ofC. bescii. These results refine the known biodiversity ofCaldicellulosiruptorand indicate that microcrystalline cellulose degradation at temperatures above 70°C, based on current information, is limited to certain members of this genus that produce GH48 domain-containing enzymes.IMPORTANCEThe genusCaldicellulosiruptorcontains the most thermophilic bacteria capable of lignocellulose deconstruction, which are promising candidates for consolidated bioprocessing for the production of biofuels and bio-based chemicals. The focus here is on the extant capability of this genus for plant biomass degradation and the extent to which this can be inferred from the core and pangenomes, based on analysis of 13 species and metagenomic sequence information from environmental samples. Key to microcrystalline hydrolysis is the content of the glucan degradation locus (GDL), a set of genes encoding glycoside hydrolases (GHs), several of which have GH48 and family 3 carbohydrate binding module domains, that function as primary cellulases. Resolving the relationship between the GDL and lignocellulose degradation will inform efforts to identify more prolific members of the genus and to develop metabolic engineering strategies to improve this characteristic.

scholarly journals A Multiomic Approach to Understand How Pleurotus eryngii Transforms Non-Woody Lignocellulosic Material

2021 ◽  
Vol 7 (6) ◽  
pp. 426
Author(s):  
Ander Peña ◽  
Rashid Babiker ◽  
Delphine Chaduli ◽  
Anna Lipzen ◽  
Mei Wang ◽  
...  
Keyword(s):  
Extracellular Enzymes ◽  
Fungal Growth ◽  
2D Nmr ◽  
Pleurotus Eryngii ◽  
Glycoside Hydrolases ◽  
Oxidative Enzymes ◽  
Initial Growth ◽  
Lignocellulosic Material ◽  
Polysaccharide Lyases ◽  
Polysaccharide Monooxygenases

Pleurotus eryngii is a grassland-inhabiting fungus of biotechnological interest due to its ability to colonize non-woody lignocellulosic material. Genomic, transcriptomic, exoproteomic, and metabolomic analyses were combined to explain the enzymatic aspects underlaying wheat–straw transformation. Up-regulated and constitutive glycoside–hydrolases, polysaccharide–lyases, and carbohydrate–esterases active on polysaccharides, laccases active on lignin, and a surprisingly high amount of constitutive/inducible aryl–alcohol oxidases (AAOs) constituted the suite of extracellular enzymes at early fungal growth. Higher enzyme diversity and abundance characterized the longer-term growth, with an array of oxidoreductases involved in depolymerization of both cellulose and lignin, which were often up-regulated since initial growth. These oxidative enzymes included lytic polysaccharide monooxygenases (LPMOs) acting on crystalline polysaccharides, cellobiose dehydrogenase involved in LPMO activation, and ligninolytic peroxidases (mainly manganese-oxidizing peroxidases), together with highly abundant H2O2-producing AAOs. Interestingly, some of the most relevant enzymes acting on polysaccharides were appended to a cellulose-binding module. This is potentially related to the non-woody habitat of P. eryngii (in contrast to the wood habitat of many basidiomycetes). Additionally, insights into the intracellular catabolism of aromatic compounds, which is a neglected area of study in lignin degradation by basidiomycetes, were also provided. The multiomic approach reveals that although non-woody decay does not result in dramatic modifications, as revealed by detailed 2D-NMR and other analyses, it implies activation of the complete set of hydrolytic and oxidative enzymes characterizing lignocellulose-decaying basidiomycetes.

scholarly journals Cross-Feeding of a Toxic Metabolite in a Synthetic Lignocellulose-Degrading Microbial Community

2021 ◽  
Vol 9 (2) ◽  
pp. 321
Author(s):  
Jessica A. Lee ◽  
Alyssa C. Baugh ◽  
Nicholas J. Shevalier ◽  
Brandi Strand ◽  
Sergey Stolyar ◽  
...  
Keyword(s):  
Resource Competition ◽  
Plant Biomass ◽  
Lignocellulose Degradation ◽  
Toxic Metabolite ◽  
Growth Requirements ◽  
Mutualistic Interaction ◽  
Metabolite Exchange ◽  
Future Work ◽  
Biomass Transformation ◽  
Complex Organic

The recalcitrance of complex organic polymers such as lignocellulose is one of the major obstacles to sustainable energy production from plant biomass, and the generation of toxic intermediates can negatively impact the efficiency of microbial lignocellulose degradation. Here, we describe the development of a model microbial consortium for studying lignocellulose degradation, with the specific goal of mitigating the production of the toxin formaldehyde during the breakdown of methoxylated aromatic compounds. Included are Pseudomonas putida, a lignin degrader; Cellulomonas fimi, a cellulose degrader; and sometimes Yarrowia lipolytica, an oleaginous yeast. Unique to our system is the inclusion of Methylorubrum extorquens, a methylotroph capable of using formaldehyde for growth. We developed a defined minimal “Model Lignocellulose” growth medium for reproducible coculture experiments. We demonstrated that the formaldehyde produced by P. putida growing on vanillic acid can exceed the minimum inhibitory concentration for C. fimi, and, furthermore, that the presence of M. extorquens lowers those concentrations. We also uncovered unexpected ecological dynamics, including resource competition, and interspecies differences in growth requirements and toxin sensitivities. Finally, we introduced the possibility for a mutualistic interaction between C. fimi and M. extorquens through metabolite exchange. This study lays the foundation to enable future work incorporating metabolomic analysis and modeling, genetic engineering, and laboratory evolution, on a model system that is appropriate both for fundamental eco-evolutionary studies and for the optimization of efficiency and yield in microbially-mediated biomass transformation.

scholarly journals Properties of Thermobifida fusca peroxidase Tfu-1649 and its combined synergistic effects with xylanase on lignocellulose degradation

BioResources ◽  
2020 ◽  
Vol 16 (1) ◽  
pp. 942-953
Author(s):  
Wan-Yu Liao ◽  
Yu-Chun Huang ◽  
Wei-Lin Chen ◽  
Cheng-Yu Chen ◽  
Chao-Hsun Yang
Keyword(s):  
Reducing Sugars ◽  
Plant Biomass ◽  
Synergistic Effects ◽  
Culture Broth ◽  
Lignocellulose Degradation ◽  
Total Phenolic ◽  
Thermobifida Fusca ◽  
Zizania Latifolia ◽  
Thermophilic Actinomycetes ◽  
Biomass Decomposition

Lignocelluloses are comprised of cellulose, hemicellulose, and lignins, which constitute plant biomass. Since peroxidases can degrade lignins, the authors examined peroxidase Tfu-1649, which is secreted from the thermophilic actinomycetes, Thermobifida fusca BCRC 19214. After cultivating for 48 h, the culture broth accumulated 43.66 U/mL of peroxidase activity. The treatment of four types of lignocellulolytic byproducts, i.e., bagasse, corncob, pin sawdust, and Zizania latifolia Turcz husk, with Tfu-1649 alone increased the total phenolic compounds, with limited reducing sugars, but treatment with xylanase, Tfu-11, and peroxidase Tfu-1649 showed synergistic effects. Hence, the co-operative degradation of lignocelluloses by both peroxidase and xylanase could contribute to biomass decomposition and further applications in the agricultural and environmental industries.

scholarly journals Soil-Derived Microbial Consortia Enriched with Different Plant Biomass Reveal Distinct Players Acting in Lignocellulose Degradation

2015 ◽  
Vol 71 (3) ◽  
pp. 616-627 ◽  
Author(s):  
Maria Julia de Lima Brossi ◽  
Diego Javier Jiménez ◽  
Larisa Cortes-Tolalpa ◽  
Jan Dirk van Elsas
Keyword(s):  
Plant Biomass ◽  
Microbial Consortia ◽  
Lignocellulose Degradation

scholarly journals Genomic and Transcriptome Analyses of a Thermophilic Bacterium Geobacillus stearothermophilus B5 Isolated from Compost Reveal Its Enzymatic Basis for Lignocellulose Degradation

2020 ◽  
Vol 8 (9) ◽  
pp. 1357
Author(s):  
Mengmeng Wang ◽  
Jiaxi Miao ◽  
Xuanqing Wang ◽  
Tuo Li ◽  
Han Zhu ◽  
...  
Keyword(s):  
Thermophilic Bacterium ◽  
Glycoside Hydrolases ◽  
Lignocellulose Degradation ◽  
Geobacillus Stearothermophilus ◽  
Carbohydrate Active Enzymes ◽  
Transcriptional Responses ◽  
Protein Coding ◽  
Clusters Of Orthologous Groups ◽  
Different Temperatures ◽  
Shock Proteins

A lignocellulose-degrading strain isolated from thermophilic compost was identified as Geobacillus stearothermophilus B5, and found able to secrete considerable amounts of enzymes at optimal temperature (60 °C) and pH (7.5). One circular contig of 3.37 Mbp was assembled from raw data, and 3371 protein-coding genes were predicted. Clusters of orthologous groups (COG) analysis revealed various genes with functions in polymeric substrate degradation, especially for Carbohydrate Active enZymes (CAZymes), such as glycoside hydrolases (GHs) and glycosyl transferases (GTs). Furthermore, the transcriptional responses of B5 at different temperatures—with rice straw provided as the sole carbon source—were analyzed. The results revealed that B5 could resist high temperature by upregulating heat shock proteins (HSPs), enhancing protein synthesis, and decreasing carbon catabolism. Briefly, B5 possesses the ability of lignocellulose degradation, and might be considered a potential inoculant for improving composting efficiency.

scholarly journals ABC Transporters Required for Hexose Uptake byClostridium phytofermentans

2019 ◽  
Vol 201 (15) ◽  
Author(s):  
Tristan Cerisy ◽  
Alba Iglesias ◽  
William Rostain ◽  
Magali Boutard ◽  
Christine Pelle ◽  
...  
Keyword(s):  
Abc Transporters ◽  
Extracellular Enzymes ◽  
Plant Biomass ◽  
Carbon Sources ◽  
Anaerobic Bacteria ◽  
Model Organism ◽  
Industrial Transformation ◽  
Content Type ◽  
Plant Matter

ABSTRACTThe mechanisms by which bacteria uptake solutes across the cell membrane broadly impact their cellular energetics. Here, we use functional genomic, genetic, and biophysical approaches to reveal howClostridium(Lachnoclostridium)phytofermentans, a model bacterium that ferments lignocellulosic biomass, uptakes plant hexoses using highly specific, nonredundant ATP-binding cassette (ABC) transporters. We analyze the transcription patterns of its 173 annotated sugar transporter genes to find those upregulated on specific carbon sources. Inactivation of these genes reveals that individual ABC transporters are required for uptake of hexoses and hexo-oligosaccharides and that distinct ABC transporters are used for oligosaccharides versus their constituent monomers. The thermodynamics of sugar binding shows that substrate specificity of these transporters is encoded by the extracellular solute-binding subunit. As sugars are not phosphorylated during ABC transport, we identify intracellular hexokinases based onin vitroactivities. These mechanisms used byClostridiato uptake plant hexoses are key to understanding soil and intestinal microbiomes and to engineer strains for industrial transformation of lignocellulose.IMPORTANCEPlant-fermentingClostridiaare anaerobic bacteria that recycle plant matter in soil and promote human health by fermenting dietary fiber in the intestine.Clostridiadegrade plant biomass using extracellular enzymes and then uptake the liberated sugars for fermentation. The main sugars in plant biomass are hexoses, and here, we identify how hexoses are taken in to the cell by the model organismClostridium phytofermentans. We show that this bacterium uptakes hexoses using a set of highly specific, nonredundant ABC transporters. Once in the cell, the hexoses are phosphorylated by intracellular hexokinases. This study provides insight into the functioning of abundant members of soil and intestinal microbiomes and identifies gene targets to engineer strains for industrial lignocellulosic fermentation.

scholarly journals A structural and kinetic survey of GH5_4 endoglucanases reveals determinants of broad substrate specificity and opportunities for biomass hydrolysis

2020 ◽  
Vol 295 (51) ◽  
pp. 17752-17769
Author(s):  
Evan M. Glasgow ◽  
Elias I. Kemna ◽  
Craig A. Bingman ◽  
Nicole Ing ◽  
Kai Deng ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Protein Design ◽  
Plant Biomass ◽  
Catalytic Efficiency ◽  
Structural Features ◽  
Glycoside Hydrolases ◽  
Design Strategies ◽  
Biomass Hydrolysis ◽  
Diverse Range ◽  
Broad Specificity

Broad-specificity glycoside hydrolases (GHs) contribute to plant biomass hydrolysis by degrading a diverse range of polysaccharides, making them useful catalysts for renewable energy and biocommodity production. Discovery of new GHs with improved kinetic parameters or more tolerant substrate-binding sites could increase the efficiency of renewable bioenergy production even further. GH5 has over 50 subfamilies exhibiting selectivities for reaction with β-(1,4)–linked oligo- and polysaccharides. Among these, subfamily 4 (GH5_4) contains numerous broad-selectivity endoglucanases that hydrolyze cellulose, xyloglucan, and mixed-linkage glucans. We previously surveyed the whole subfamily and found over 100 new broad-specificity endoglucanases, although the structural origins of broad specificity remained unclear. A mechanistic understanding of GH5_4 substrate specificity would help inform the best protein design strategies and the most appropriate industrial application of broad-specificity endoglucanases. Here we report structures of 10 new GH5_4 enzymes from cellulolytic microbes and characterize their substrate selectivity using normalized reducing sugar assays and MS. We found that GH5_4 enzymes have the highest catalytic efficiency for hydrolysis of xyloglucan, glucomannan, and soluble β-glucans, with opportunistic secondary reactions on cellulose, mannan, and xylan. The positions of key aromatic residues determine the overall reaction rate and breadth of substrate tolerance, and they contribute to differences in oligosaccharide cleavage patterns. Our new composite model identifies several critical structural features that confer broad specificity and may be readily engineered into existing industrial enzymes. We demonstrate that GH5_4 endoglucanases can have broad specificity without sacrificing high activity, making them a valuable addition to the biomass deconstruction toolset.

Improved lignocellulose degradation efficiency based on Fenton pretreatment during rice straw composting

2019 ◽  
Vol 294 ◽  
pp. 122132 ◽  
Author(s):  
Di Wu ◽  
Zimin Wei ◽  
Yue Zhao ◽  
Xinyu Zhao ◽  
Taha Ahmed Mohamed ◽  
...  
Keyword(s):  
Rice Straw ◽  
Degradation Efficiency ◽  
Lignocellulose Degradation

Enhanced degradation of ammonium-pretreated wheat straw by lignocellulolytic Streptomyces spp.

1992 ◽  
Vol 38 (10) ◽  
pp. 1022-1025 ◽  
Author(s):  
Marina Basaglia ◽  
Giuseppe Concheri ◽  
Stefano Cardinali ◽  
Maria B. Pasti-Grigsby ◽  
Marco P. Nuti
Keyword(s):  
Weight Loss ◽  
Wheat Straw ◽  
Lignin Degradation ◽  
Cellulose Degradation ◽  
Lignocellulose Degradation ◽  
Cellulose Content ◽  
Streptomyces Spp ◽  
Initial Substrate ◽  
Streptomyces Chromofuscus ◽  
Pretreated Wheat Straw

Eleven actinomycetes, isolated from the gut of worker termites (Macrotermes, Armitermes, Microcerotermes, Odontotermes), were identified as Streptomyces chromofuscus, S. chromogenus, S. diastaticus, and S. rochei. Their ability to grow on natural lignocellulosic substrates was tested in solid state fermentation experiments using wheat straw (C/N = 49.8) as a sole carbon source. Weight loss was 4.7–20.9% of the initial substrate, after 5 weeks at 30 °C; lignin and cellulose content decreased 2.0–16.1 and 3.5–32.9%, respectively. When the 11 Streptomyces were grown on wheat straw pretreated with (NH4)HCO3 (C/N = 28.2), weight loss was 9.3–29.9% of the initial substrate, indicating an overall enhancement of lignocellulose degradation. Weight, lignin, and cellulose losses were enhanced when S. chromofuscus (strain A2 and A11) and S. rochei A4 were grown on pretreated wheat straw instead of the untreated substrate. With S. rochei A10 the weight loss and lignin degradation were enhanced, while cellulolysis was slightly depressed. Weight loss and cellulose degradation were both enhanced when the remaining strains were grown on pretreated wheat straw. In this case, lignin degradation was depressed (S. chromofuscus A6 and A8, S. diastaticus A12, S. rochei A14) or remained essentially the same (S. diastaticus A3 and S. chromogenus A7). Key words: Streptomyces, wheat straw, degradation, lignin, cellulose.

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