Dataset Information

Bioethanol Production from Brewers Spent Grains Using a Fungal Consolidated Bioprocessing (CBP) Approach.

ABSTRACT: Production of bioethanol from brewers spent grains (BSG) using consolidated bioprocessing (CBP) is reported. Each CBP system consists of a primary filamentous fungal species, which secretes the enzymes required to deconstruct biomass, paired with a secondary yeast species to ferment liberated sugars to ethanol. Interestingly, although several pairings of fungi were investigated, the sake fermentation system (A. oryzae and S. cerevisiae NCYC479) was found to yield the highest concentrations of ethanol (37 g/L of ethanol within 10 days). On this basis, 1 t of BSG (dry weight) would yield 94 kg of ethanol using 36 hL of water in the process. QRT-PCR analysis of selected carbohydrate degrading (CAZy) genes expressed by A. oryzae in the BSG sake system showed that hemicellulose was deconstructed first, followed by cellulose. One drawback of the CBP approach is lower ethanol productivity rates; however, it requires low energy and water inputs, and hence is worthy of further investigation and optimisation.

SUBMITTER: Wilkinson S

PROVIDER: S-EPMC7114960 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Bioethanol Production from Brewers Spent Grains Using a Fungal Consolidated Bioprocessing (CBP) Approach.

Wilkinson Stuart S Smart Katherine A KA James Sue S Cook David J DJ

Bioenergy research 20160808 1

Production of bioethanol from brewers spent grains (BSG) using consolidated bioprocessing (CBP) is reported. Each CBP system consists of a primary filamentous fungal species, which secretes the enzymes required to deconstruct biomass, paired with a secondary yeast species to ferment liberated sugars to ethanol. Interestingly, although several pairings of fungi were investigated, the sake fermentation system (<i>A. oryzae</i> and <i>S. cerevisiae</i> NCYC479) was found to yield the highest concen ...[more]

PMID: 32269706

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Project description:Natural yeast with superior fermentative traits can serve as a platform for the development of recombinant strains that can be used to improve the sustainability of bioethanol production from starch. This process will benefit from a consolidated bioprocessing (CBP) approach where an engineered strain producing amylases directly converts starch into ethanol. The yeast Saccharomyces cerevisiae L20, previously selected as outperforming the benchmark yeast Ethanol Red, was here subjected to a comparative genomic investigation using a dataset of industrial S. cerevisiae strains. Along with Ethanol Red, strain L20 was then engineered for the expression of α-amylase amyA and glucoamylase glaA genes from Aspergillus tubingensis by employing two different approaches (delta integration and CRISPR/Cas9). A correlation between the number of integrated copies and the hydrolytic abilities of the recombinants was investigated. L20 demonstrated important traits for the construction of a proficient CBP yeast. Despite showing a close relatedness to commercial wine yeast and the benchmark Ethanol Red, a unique profile of gene copy number variations (CNVs) was found in L20, mainly encoding membrane transporters and secretion pathway proteins but also the fermentative metabolism. Moreover, the genome annotation disclosed seven open reading frames (ORFs) in L20 that are absent in the reference S288C genome. Genome engineering was successfully implemented for amylase production. However, with equal amylase gene copies, L20 proved its proficiency as a good enzyme secretor by exhibiting a markedly higher amylolytic activity than Ethanol Red, in compliance to the findings of the genomic exploration. The recombinant L20 dT8 exhibited the highest amylolytic activity and produced more than 4 g/L of ethanol from 2% starch in a CBP setting without the addition of supplementary enzymes. Based on the performance of this strain, an amylase/glucoamylase ratio of 1:2.5 was suggested as baseline for further improvement of the CBP ability. Overall, L20 showed important traits for the future construction of a proficient CBP yeast. As such, this work shows that natural S. cerevisiae strains can be used for the expression of foreign secreted enzymes, paving the way to strain improvement for the starch-to-bioethanol route.

Project description:BACKGROUND: The recalcitrant nature of hemicellulosic materials and the high cost in depolymerization are the primary obstacles preventing the use of xylan as feedstock for fuel and chemical production. Consolidated bioprocessing, incorporating enzyme-generating, biomass-degrading and bioproduct-producing capabilities into a single microorganism, could potentially avoid the cost of the dedicated enzyme generation in the process of xylan utilization. In this study, we engineered Escherichia coli strains capable of exporting three hemicellulases to the broth for the succinate production directly from beechwood xylan. RESULTS: Xylanases were extracellular environment-directed by fusing with OsmY. Subsequently, twelve variant OsmY fused endoxylanase-xylosidase combinations were characterized and tested. The combination of XynC-A from Fibrobacter succinogenes S85 and XyloA from Fusarium graminearum which appeared to have optimal enzymatic properties was identified as the best choice for xylan hydrolysis (0.18 ± 0.01 g/l protein in the broth with endoxylanase activity of 12.14 ± 0.34 U/mg protein and xylosidase activity of 92 ± 3 mU/mg protein at 8 h after induction). Further improvements of hemicellulases secretion were investigated by lpp deletion, dsbA overexpression and expression level optimization. With co-expression of ?-arabinofuranosidase, the engineered E. coli could hydrolyze beechwood xylan to pentose monosaccharides. The hemicellulolytic capacity was further integrated with a succinate-producing strain to demonstrate the production of succinate directly from xylan without externally supplied hydrolases and any other organic nutrient. The resulting E. coli Z6373 was able to produce 0.37 g/g succinate from xylan anaerobically equivalent to 76% of that from xylan acid hydrolysates. CONCLUSIONS: This report represents a promising step towards the goal of hemicellulosic chemical production. This engineered E. coli expressing and secreting three hemicellulases demonstrated a considerable succinate production on the released monosaccharides from xylan. The ability to use lower-cost crude feedstock will make biological succinate production more economically attractive.

Dataset Information

Bioethanol Production from Brewers Spent Grains Using a Fungal Consolidated Bioprocessing (CBP) Approach.

Publications

Bioethanol Production from Brewers Spent Grains Using a Fungal Consolidated Bioprocessing (CBP) Approach.

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