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Hydrogen sulfide (H2S) is a crucial signaling molecule in plants. Recent studies have shown that H2S plays an equally important role as nitric oxide (NO) and hydrogen peroxide (H2O2) in plant signaling. Previous studies have demonstrated the involvement of H2S in regulating drought and other stressful environmental conditions, but the exact downstream molecular mechanisms activated by the H2S signaling molecule remain unclear. In this study, we conducted a comprehensive genome-wide transcriptomic analysis of both wild type (WT) and double mutant (lcd/des1). Arabidopsis thaliana plants were exposed to 40% polyethylene glycol (PEG) to induce drought stress and 20 µM sodium hydrosulfide (NaHS). The resulting transcriptome data were analyzed for differentially significant genes and their statistical enrichments in the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The results indicated significant upregulation of genes related to photosynthesis, carbon fixation, plant secondary metabolite biosynthesis, inositol and phosphatidylinositol signaling pathways, and stress-responsive pathways in mutant plants under drought stress. Mutant plants with impaired H2S signaling mechanisms displayed greater susceptibility to drought stress compared to wild-type plants. In summary, all findings highlight the pivotal role of H2S signaling in stimulating other drought-responsive signaling pathways.

Abstract Background Lignin is an intricate phenolic polymer found in plant cell walls that has tremendous potential for being converted into value-added products with the possibility of significantly increasing the economics of bio-refineries. Although lignin in nature is bio-degradable, its biocatalytic conversion is challenging due to its stable complex structure and recalcitrance. In this context, an understanding of strain's genomics, enzymes, and degradation pathways can provide a solution for breaking down lignin to unlock the full potential of lignin as a dominant valuable bioresource. A gammaproteobacterial strain AORB19 has been isolated previously from decomposed wood based on its high laccase production. This work then focused on the detailed genomic and functional characterization of this strain based on whole genome sequencing, the identification of lignin degradation products, and the strain’s laccase production capabilities on various agro-industrial residues. Results Lignin degrading bacterial strain AORB19 was identified as Serratia quinivorans based on whole genome sequencing and core genome phylogeny. The strain comprised a total of 123 annotated CAZyme genes, including ten cellulases, four hemicellulases, five predicted carbohydrate esterase genes, and eight lignin-degrading enzyme genes. Strain AORB19 was also found to possess genes associated with metabolic pathways such as the β-ketoadipate, gentisate, anthranilate, homogentisic, and phenylacetate CoA pathways. LC–UV analysis demonstrated the presence of p-hydroxybenzaldehyde and vanillin in the culture media which constitutes potent biosignatures indicating the strain’s capability to degrade lignin. Finally, the study evaluated the laccase production of Serratia AORB19 grown with various industrial raw materials, with the highest activity detected on flax seed meal (257.71 U/L), followed by pea hull (230.11 U/L), canola meal (209.56 U/L), okara (187.67 U/L), and barley malt sprouts (169.27 U/L). Conclusions The whole genome analysis of Serratia quinivorans AORB19, elucidated a repertoire of genes, pathways and enzymes vital for lignin degradation that widens the understanding of ligninolytic metabolism among bacterial lignin degraders. The LC-UV analysis of the lignin degradation products coupled with the ability of S. quinivorans AORB19 to produce laccase on diverse agro-industrial residues underscores its versatility and its potential to contribute to the economic viability of bio-refineries.

According to the society of assisted reproductive technologies (SART), the top fertility clinics in North America have reported overall pregnancy rates of up to 50% in good prognosis patients. Although new techniques in advanced reproductive technologies have begun to emerge in the twenty-first century, including time-lapse monitoring of embryo development, analysis of metabolites in embryo culture media, and the use of artificial intelligence in morphometric embryo grading, currently chromosomal status obtained via pre-implantation genetic testing for aneuploidy (PGT-A) appears to be the most definitive in evaluating embryonic potential. Yet still, in most clinics, approximately 30% of euploid or chromosomally normal embryos fail to achieve a viable pregnancy resulting in stress and anxiety for patients, repeat cycles, and increased cost of treatment. Recent literature suggests that blastocysts with an increased mitochondrial DNA (mtDNA) ratio have greatly reduced implantation potential, however, there exists controversial studies which do not report a relationship. This review summarizes the current literature on utilizing mitochondrial DNA as a possible biomarker for embryo implantation, highlights the reasons for inconsistencies in some studies and suggests possible improvements to the study design of future mtDNA ratio research.

The present work demonstrated the potential of different agro-waste mixtures to produce ethanol. The forest soil bacterium (Streptomyces sp.) was exploited for the saccharification of the agro-waste mixture formulated by extreme vertices mixture design and the hydrolysate produced by saccharification was used for fermentation. The best formulation contained 43.33% orange peel, 33.33 % pumpkin pulp + seeds, and 23.33% pomegranate peel which exhibited significantly high reducing sugar (22.36 ± 0.54 mg/g dry weight) among all other mixtures. The hydrolysate of this mixture when supplemented with 2% w/v fructose produced a maximum of 7.86 ± 0.08% v/v ethanol by the yeast isolated from the brewer’s spent grains. Thus, easily available waste could be a promising source for yeast isolation and feedstock for ethanol production. Further, this study aids to reduce the risk of health, and environmental pollution, and developing the economy.

Glucose isomerase (GI) is an enzyme with high potential applications. Characterization of GI producing bacteria with interesting properties from an industrial point of view is essential. Bacillus sp., Paenarthrobacter sp., Chryseobacterium sp., Hymenobacter sp., Mycobacterium sp., and Stenotrophomonas sp. were isolated from soil samples. Optimization of enzyme production yield was investigated in various fermentation conditions using response surface methodology. All isolates exhibited maximum GI activity at 40 °C, pH 6–8 after 4 days of incubation. A mixture of peptone/yeast extract or tryptone/peptone enhanced higher enzyme production. The same trend was observed in fermentation medium containing 1% xylose or 2%–2.5% wheat straw. This study advanced the knowledge of these bacterial isolates in promoting wheat straw as feedstock for the bio-based industry.

Microplastics have become a global environmental concern due to their ubiquitous presence and persistence in the environment and have been identified as a major pollutant in aquatic environments. This study aimed to evaluate the effect of various concentrations of polyethylene terephthalate (PET) microplastic (25–200 mg/L) on the growth, chlorophyll content, and toxicity of Scenedesmus sp. from aquatic and terrestrial habitats over a period of 24 days. Our results showed that microplastics with higher concentration (200 mg/L) have a significantly higher inhibitory effect. Also, higher concentration of extracellular hydrogen peroxide (H2O2) extracellular polymeric substance (EPS) were found on microalgae exposed to the microplastic. Further, Scanning Electron Microscope (SEM) images revealed that microalgae attached to microplastic surfaces and formed hetero-aggregation. Overall, our study provided valuable information for understanding the complex effects of microplastics on microalgae, particularly in comparing the differential effects on aquatic and terrestrial Scenedesmus sp.

High throughput screening approaches can significantly speed up the identification of novel enzymes from natural microbial consortiums. A two-step high throughput screening process was proposed and explored to screen lignin-degrading microorganisms. By employing this modified culture enrichment method and screening based on enzyme activity, a total of 82 bacterial and 46 fungal strains were isolated from fifty decayed wood samples (100 liquid cultures) collected from the banks of the Ottawa River in Canada. Among them, ten bacterial and five fungal strains were selected and identified based on their high laccase activities by 16S rDNA and ITS gene sequencing, respectively. The study identified bacterial strains from various genera including Serratia, Enterobacter, Raoultella, and Bacillus, along with fungal counterparts including Mucor, Trametes, Conifera and Aspergillus. Moreover, Aspergillus sydowii (AORF21), Mucor sp. (AORF43), Trametes versicolor (AORF3) and Enterobacter sp. (AORB55) exhibited xylanase and β- glucanase activities in addition to laccase production. The proposed approach allowed for the quick identification of promising consortia and enhanced the chance of isolating desired strains based on desired enzyme activities. This method is not limited to lignocellulose and lignin-degrading microorganisms but can be applied to identify novel microbial strains and enzymes from different natural samples.

High fructose corn syrup has been industrially produced by converting glucose to fructose by glucose isomerases, tetrameric metalloenzymes widely used in industrial biocatalysis. Advances in enzyme engineering and commercial production of glucose isomerase have paved the way to explore more efficient variants of these enzymes. The 5-hydroxymethylfurfural can be produced from high fructose corn syrup catalytic dehydration, and it can be further converted into various furanic compounds chemically or biologically for various industrial applications as a promising platform chemical. Although the chemical conversion of 5-hydroxymethylfurfural into furanic compounds has been extensively investigated in recent years, bioconversion has shown promise for its mild conditions due to the harsh chemical reaction conditions. This review discusses protein engineering potential for improving glucose isomerase production and recent advancements in bioconversion of 5-hydroxymethylfurfural into value-added furanic derivatives. It suggests biological strategies for the industrial transformation of 5-hydroxymethylfurfural.

Abstract Microbial weathering processes can significantly promote soil properties and reduce rock-to-soil ratio. Some soil-inhabiting bacteria exhibit efficient rock-dissolution abilities by releasing organic acids and other chemical elements from the silicate rocks. However, our understanding of the molecular mechanisms involved during bacterial rock-dissolution is still limited. In this study, we performed silicate rock-dissolution experiments on a Pseudomonas sp. NLX-4 strain isolated from an over-exploited mining site. The results revealed that Pseudomonas sp. NLX-4 strain efficiently accelerates the dissolution of silicate rocks by secreting amino acids, exopolysaccharides, and organic acids. Through employing genome and transcriptome sequencing (RNA-seq), we identified the major regulatory genes. Specifically, 15 differentially expressed genes (DEGs) encoding for siderophore transport, EPS and amino acids synthesis, organic acids metabolism, and bacterial resistance to adverse environmental conditions were highly up-regulated in silicate rock cultures of NLX-4 strain. Our study reports a potential bacterial based approach for improving the ecological restoration of over-exploited rock mining sites. Graphical Abstract

The present study identified the pectinase-producing bacterium isolated from the contaminated broth as Bacillus sp. on 16S rDNA sequence analysis. The bacterium illustrated water-like droplets on the colony grown on the Sabouraud dextrose agar plate. It also exhibited multi-enzymes activities, such as pectinase, polygalacturonase, xylanase, and cellulase by using various agro-wastes as low-cost substrates. The orange peel was observed to be the best substrate among the agro-wastes used for maximum multi-enzymes (pectinase, polygalacturonase, xylanase, and cellulase). However, the bacterium demonstrated its capability to produce different enzymes according to the different substrates/agro-wastes used. The Plackett–Burman design was used to determine the essential influencing factors, while the Box Behnken design response surface methodology was for optimizing cultural conditions. At their optimal conditions (40°C incubation temperature, 24 h of incubation period, 1% w/v orange peel, and 2% v/v inoculum volume), the bacterium exhibited the maximum pectinase (9.49 ± 1.25 U/ml) and xylanase (16.27 ± 0.52 U/ml) activities. Furthermore, the study explored the ability of the bacterium to produce bacterial lipids and observed about 25% bacterial lipid content on a dry weight basis. Therefore, the bacterium is a good candidate for producing important multi-enzymes and subsequent agro-waste degradation controlling the environment, and facilitating waste management. Also, the bacterium can be a potential feedstock in producing renewable biofuel.

Abstract The biological pretreatment for the enzymatic hydrolysis of lignocellulosic biomasses depends exclusively on the effective pretreatment process. Herein, we report a significant enhancement of enzymatic saccharification obtained with corn stover using a bacterial strain Bacillus sp. P3. The hemicellulose removal from corn stover by the strain Bacillus sp. P3 was evaluated for enhancing subsequent enzymatic hydrolysis. Therefore, our study revealed that an alkaline-resistant xylanase as well as other enzymes produced by Bacillus sp. P3 in fermentation broth led to a substantially enhanced hemicellulose removal rate from corn stover within pH 9.36–9.68. However, after a 20-day pretreatment of corn stover by the strain P3, the glucan content was increased by 51% and the xylan content was decreased by 35%. After 72 h of saccharification using 20 U/g of commercial cellulase, the yield of reducing sugar released from 20-day pretreated corn stover was increased by 56% in comparison to the untreated corn stover. Therefore, the use of the strain P3 could be a promising approach to pretreat corn stover for enhancing the enzymatic hydrolysis process of industrial bioenergy productions.

Abstract Background Vanillin is one of the important phenolic inhibitors in Saccharomyces cerevisiae for bioconversion of lignocellulosic materials and has been reported to inhibit the translation process in cells. In our previous studies, it was confirmed that the deletion of the transcription factor gene YRR1 enhanced vanillin resistance by promoting some translation-related processes at the transcription level. In this work, we investigated the effects of proteomic changes upon induction of vanillin stress and deletion of YRR1 to provide unique perspectives from a transcriptome analysis for comprehending the mechanisms of YRR1 deletion in the protective response of yeast to vanillin. Results In wild-type cells, vanillin reduced two dozens of ribosomal proteins contents while upregulated proteins involved in glycolysis, oxidative phosphorylation, and the pentose phosphate pathway in cells. The ratios of NADPH/NADP+ and NADH/NAD+ were increased when cells responded to vanillin stress. The differentially expressed proteins perturbed by YRR1 deletion were much more abundant than and showed no overlaps with transcriptome changes, indicating that Yrr1 affects the synthesis of certain proteins. Forty-eight of 112 upregulated proteins were involved in the stress response, translational and transcriptional regulation. YRR1 deletion increased the expression of HAA1-encoding transcriptional activator, TMA17-encoding proteasome assembly chaperone and MBF1-encoding coactivator at the protein level, as confirmed by ELISA. Cultivation data showed that the overexpression of HAA1 and TMA17 enhanced resistance to vanillin in S. cerevisiae. Conclusions Cells conserve energy by decreasing the content of ribosomal proteins, producing more energy and NAD(P)H for survival in response to vanillin stress. Yrr1 improved vanillin resistance by increasing the protein quantities of Haa1, Tma17 and Mbf1. These results showed the response of S. cerevisiae to vanillin and how YRR1 deletion increases vanillin resistance at the protein level. These findings may advance our knowledge of how YRR1 deletion protects yeast from vanillin stress and offer novel targets for genetic engineering of designing inhibitor-resistant ethanologenic yeast strains.

Natural biodegradation processes hold promises for the conversion of agro-industrial lignocellulosic biomaterials into biofuels and fine chemicals through lignin-degrading enzymes. The high cost and low stability of these enzymes remain a significant challenge to economic lignocellulosic biomass conversion. Wood-degrading microorganisms are a great source for novel enzyme discoveries. In this study, the decomposed wood samples were screened, and a promising γ-proteobacterial strain that naturally secreted a significant amount of laccase enzyme was isolated and identified as Serratia proteamaculans AORB19 based on its phenotypic and genotypic characteristics. The laccase activities in culture medium of strain AORB19 were confirmed both qualitatively and quantitatively. Significant cultural parameters for laccase production under submerged conditions were identified following a one-factor-at-a-time (OFAT) methodology: temperature 30°C, pH 9, yeast extract (2 g/l), Li+, Cu2+, Ca2+, and Mn2+ (0.5 mM), and acetone (5%). Under the selected conditions, a 6-fold increase (73.3 U/L) in laccase production was achieved when compared with the initial culturing conditions (12.18 U/L). Furthermore, laccase production was enhanced under alkaline and mesophilic growth conditions in the presence of metal ions and organic solvents. The results of the study suggest the promising potential of the identified strain and its enzymes in the valorization of lignocellulosic wastes. Further optimization of culturing conditions to enhance the AORB19 strain laccase secretion, identification and characterization of the purified enzyme, and heterologous expression of the specific enzyme may lead to practical industrial and environmental applications.

Summary Saccharomyces cerevisiae is the preferred source of RNA derivatives, which are widely used as supplements for foods and pharmaceuticals. As the most abundant RNAs, the ribosomal RNAs (rRNAs) transcribed by RNA polymerase I (Pol I) have no 5′ caps, thus cannot be translated to proteins. To screen high‐nucleic‐acid content yeasts more efficiently, a cap‐independent protein expression system mediated by Pol I has been designed and established to monitor the regulatory changes of rRNA synthesis by observing the variation in the reporter genes expression. The elements including Pol I‐recognized rDNA promoter, the internal ribosome entry site from cricket paralytic virus which can recruit ribosomes internally, reporter genes (URA3 and yEGFP3), oligo‐dT and an rDNA terminator were ligated to a yeast episomal plasmid. This system based on the URA3 gene worked well by observing the growth phenotype and did not require the disruption of cap‐dependent initiation factors. The fluorescence intensity of strains expressing the yEGFP3 gene increased and drifted after mutagenesis. Combined with flow cytometry, cells with higher GFP level were sorted out. A strain showed 58% improvement in RNA content and exhibited no sequence alteration in the whole expression cassette introduced. This study provides a novel strategy for breeding high‐nucleic‐acid content yeasts.

Brown-rot fungi are rapid holocellulose degraders and are the most predominant degraders of coniferous wood products in North America. Brown-rot fungi degrades wood by both enzymatic (plant biomass degrading carbohydrate active enzymes-CAZymes) and non-enzymatic systems (Fenton’s reaction) mechanisms. Identifying the genes and molecular mechanisms involved in Fenton’s reaction would significantly improve our understanding about brown-rot decay. Our present study identifies the common gene expression patterns involved in brown rot decay by performing metadata analysis of fungal transcriptome datasets. We have also analyzed and compared the genome-wide annotations (InterPro and CAZymes) of the selected brown rot fungi. Genes encoding for various oxidoreductases, iron homeostasis, and metabolic enzymes involved in Fenton’s mechanism were found to be significantly expressed among all the brown-rot fungal datasets. Interestingly, a higher number of hemicellulases encoding genes were differentially expressed among all the datasets, while a fewer number of cellulases and peroxidases were expressed (especially haem peroxidase and chloroperoxidase). Apart from these lignocellulose degrading enzymes genes encoding for aldo/keto reductases, 2-nitro dioxygenase, aromatic-ring dioxygenase, dienelactone hydrolase, alcohol dehydrogenase, major facilitator superfamily, cytochrome-P450 monoxygenase, cytochrome b5, and short-chain dehydrogenase were common and differentially up regulated among all the analyzed brown-rot fungal datasets.

Abstract Carbohydrate-active enzymes (CAZymes) are industrially important enzymes, which are involved in synthesis and breakdown of carbohydrates. CAZymes secreted by microorganisms especially fungi are widely used in industries. However, identifying an ideal fungal candidate is costly and time-consuming process. In this regard, we have developed a web-database “CAZymes Based Ranking of Fungi (CBRF)”, for sorting and selecting an ideal fungal candidate based on their genome-wide distribution of CAZymes. We have retrieved the complete annotated proteomic data of 443 published fungal genomes from JGI-MycoCosm web-repository, for the CBRF web-database construction. CBRF web-database was developed using open source computing programing languages such as MySQL, HTML, CSS, bootstrap, jQuery, JavaScript and Ajax frameworks. CBRF web-database sorts complete annotated list of fungi based on three selection functionalities: (a) to sort either by ascending (or) descending orders; (b) to sort the fungi based on a selected CAZy group and class; (c) to sort fungi based on their individual lignocellulolytic abilities. We have also developed a simple and basic webpage “S-CAZymes” using HTML, CSS and Java script languages. The global search functionality of S-CAZymes enables the users to understand and retrieve information about a specific carbohydrate-active enzyme and its current classification in the corresponding CAZy family. The S-CAZymes is a supporting web page which can be used in complementary with the CBRF web-database (knowing the classification of specific CAZyme in S-CAZyme and use this information further to sort fungi using CBRF web-database). The CBRF web-database and S-CAZymes webpage are hosted through Amazon® Web Services (AWS) available at http://13.58.192.177/RankEnzymes/about. We strongly believe that CBRF web-database simplifies the process of identifying a suitable fungus both in academics and industries. In future, we intend to update the CBRF web-database with the public release of new annotated fungal genomes.

Abstract In the last two decades, studies on plant biomass-degrading fungi have remarkably increased to understand and reveal the underlying molecular mechanisms responsible for their life cycle and wood-decaying abilities. Most of the plant biomass-degrading fungi reported till date belong to basidiomycota or ascomycota phyla. Thus, very few studies were conducted on fungi belonging to other divisions. Recent sequencing studies have revealed complete genomic sequences of various fungi. Our present study is focused on understanding the plant biomass-degrading potentials, by retrieving genome-wide annotations of 56 published fungi belonging to Glomeromycota, Mucoromycota, Zoopagomycota, Blastocladiomycota, Chytridiomycota, Neocallimastigomycota, Microsporidia and Cryptomycota from JGI-MycoCosm repository. We have compared and analyzed the proteomic annotations, especially CAZy, KOG, KEGG and SM clusters by separating the proteomic annotations into lignin-, cellulose-, hemicellulose-, pectin-degrading enzymes and also highlighted the KEGG, KOG molecular mechanisms responsible for the metabolism of carbohydrates (lignocellulolytic pathways of fungi), complex organic pollutants, xenobiotic compounds, biosynthesis of secondary metabolites. However, we strongly agree that studying genome-wide distributions of fungal CAZyme does not completely corresponds to its biomass-degrading ability. Thus, our present study can be used as preliminary materials for selecting ideal fungal candidate for the degradation and conversion of plant biomass components, especially carbohydrates to bioethanol and other commercially valuable products.

Summary Biomass recalcitrance is still a main challenge for the production of biofuels and high‐value products. Here, an alternative Miscanthus pretreatment method by using lignin‐degrading bacteria was developed. Six efficient Miscanthus‐degrading bacteria were first cultured to produce laccase by using 0.5% Miscanthus biomass as carbon source. After 1–5 days of incubation, the maximum laccase activities induced by Miscanthus in the six strains were ranged from 103 to 8091 U l−1. Then, the crude enzymes were directly diluted by equal volumes of citrate buffer and added Miscanthus biomass to a solid concentration at 4% (w/v). The results showed that all bacterial pretreatments significantly decreased the lignin content, especially in the presence of two laccase mediators (ABTS and HBT). The lignin removal directly correlated with increases in total sugar and glucose yields after enzymatic hydrolysis. When ABTS was used as a mediator, the best lignin‐degrading bacteria (Pseudomonas sp. AS1) can remove up to 50.1% lignin of Miscanthus by obtaining 2.2‐fold glucose yield, compared with that of untreated biomass. Therefore, this study provided an effective Miscanthus pretreatment method by using lignin‐degrading bacteria, which may be potentially used in improving enzymatic hydrolysability of biomass.

Microalgae have the potential to grow in nutrient-rich environments and have the ability to accumulate nutrients from wastewater. The nutrients in malting wastewater are ideal for microalgae cultivation. However, there is limited published work on the growth characteristics of freshwater microalgae grown in malting effluent. This study examined the potential of diluted malting effluent for the growth of freshwater green algae Chlorella sp. and Chlamydomonas sp. isolated from northern Ontario and subsequent biomass and lipid production. Under the 18:6 h light/dark cultivation cycle, the highest cell number counted (540 × 104 cell·mL−1 on day 20) and total chlorophyll content were found in 50% diluted malting effluents for Chlorella sp., whereas the 70% dilution concentration was the most productive for Chlamydomonas (386 × 104 cell·mL−1 on day 16). The total lipid content was higher in the 50% dilution concentration of malting effluent in both Chlorella sp. (maximum 20.5%–minimum 11.5% of dry weight) and Chlamydomonas sp. (max 39.3%–min 25.9% of dry weight). These results emphasize the suitability of using unsterile diluted malting effluent for microalgae cultivation.

The efficient industrial conversion of plant-derived cellulose to simple sugars and other value-added chemicals requires various highly stable and reactive enzymes. Industrial processes especially synchronous saccharification and fermentation (SSF)-based production of cellulosic bio-ethanol require enzymes that are active at lower temperatures. In this study, we have identified, characterized, and expressed the cold-adaptive endo-1,4-β-glucanase (BpEG) isolated from the Burkholderia pyrrocinia JK-SH007. The analysis of the predicted amino acid sequence indicated that BpEG belongs to GH family 8. The BpEG without the signal peptide was cloned into the expression vector pET32a and significantly expressed in Escherichia coli BL21 (DE3) competent cells. The SDS-PAGE and Western blot analysis of BpEG revealed that the recombinant BpEG was approximately 60 kDa. Purified recombinant BpEG exhibited hydrolytic activity against carboxymethyl cellulose (CMC) and phosphoric acid swollen cellulose (PASC), but not crystalline cellulose and xylan substrates. High performance, anion exchange, chromatography-pulsed amperometric detector (HPAEC-PAD) analysis of the enzymatic products obtained from depolymerization of 1,4-β-linked biopolymers of different lengths revealed an interesting cutting mechanism employed by endoglucanases. The recombinant BpEG exhibited 6.0 of optimum pH and 35°C of optimum temperature, when cultured with CMC substrate. The BpEG enzyme exhibited stable activity between pH 5.0 and 9.0 at 35°C. Interestingly, BpEG retained about 42% of its enzymatic activity at 10°C compared to its optimal temperature. This new cold-adaptive cellulase could potentially achieve synchronous saccharification and fermentation (SSF) making BpEG a promising candidate in the fields of biofuel, biorefining, food and pharmaceutical industries.

Acetyl and methyl esterifications are two major naturally found substitutions in the plant cell-wall polysaccharides. The non-cellulosic plant cell-wall polysaccharides such as pectin and hemicellulose are differentially esterified by the O-acetyl and methyl groups to cease the action of various hydrolytic enzymes secreted by different fungi and bacterial species. Thus, microorganisms have emerged with a special class of enzymes known as carbohydrate esterases (CE). The CE catalyse O-de, N-deacetylation of acetylated saccharide residues (esters or amides, where sugars play the role of alcohol/amine/acid). Carbohydrate active enzyme (CAZy) database has classified CE into 16 classes, of which hemicellulose deacetylating CE were grouped into eight classes (CE-1 to CE-7 and CE-16). Various plant biomass degrading fungi and bacteria secretes acetyl xylan esterases (AcXE); however, these enzymes exhibit varied substrate specificities. AcXE and xylanases-coupled pretreatment methods exhibit significant applications, such as enhancing animal feedstock, baking industry, production of food additives, paper and pulp, xylitol production and biorefinery-based industries, respectively. Thus, understanding the structural and functional properties of acetyl xylan esterase will significantly aid in developing the efficient AcXE with wide range of industrial applications.

Abstract Biological conversion of plant biomass into commercially valuable products is one of the highly studied subjects in the last two decades. Studies were continuously being conducted to understand and develop efficient enzymes for the breakdown and conversion of plant cell-wall components into valuable commercial products. Naturally, plant cell-wall components are differentially esterified to protect from the invading microorganisms. However, during the process of evolution, microorganisms have developed special set of enzymes to de-esterify the plant cell-wall components. Among the carbohydrate-active enzymes (CAZy), carbohydrate esterases stand first during the process of enzymatic conversion of plant biomass, as these enzymes de-esterify the plant biomass and make it accessible for the hydrolytic enzymes such as cellulases, hemicellulases, ligninolytic and pectinases. In this article, we have extensively discussed about the structural and functional properties of pectin methyl esterases, feruloyl, cinnamoyl and glucuronoyl esterases which are required for the de-esterification of pectin and lignin–carbohydrate complexes. Pectin esterases are classified among CE8, CE12, CE13 and CE15 carbohydrate esterase class of CAZy database. Whereas, lignin–carbohydrate complex de-esterifying enzymes are classified among CE1 (feruloyl esterase) and CE15 (glucuronoyl esterase) classes. Understanding the structural and functional abilities of pectin and lignin–carbohydrate esterases will significantly aid in developing efficient class of de-esterases for reducing the recalcitrant nature of plant biomass. These efficient de-esterases will have various applications including pretreatment of plant biomass, food, beverage, pulp and paper, textile, pharmaceutical and biofuel industries.

Abstract Background Xylan removal by bacterial pretreatments has been confirmed to increase the digestibility of biomass. Here, an effective xylan removal technique has been developed to enhance the digestibility of wheat straw and simultaneously produce bioflocculants by a cellulase-free xylanase-secreting strain, Pseudomonas boreopolis G22. Results The results indicated that P. boreopolis G22 is an alkaliphilic strain which can secrete abundant amounts of xylanase. This xylanase had activity levels of 2.67–1.75 U mL−1 after an incubation period of 5–25 days. The xylanase showed peak activity levels at pH 8.6, and retained more than 85% relative activity in the pH range of 7.2–9.8. After 15 days of cultivation, the hemicellulose contents of the wheat straw were significantly decreased by 32.5%, while its cellulose contents were increased by 27.3%, compared to that of the control. The maximum reducing sugars released from the 15-day-pretreated wheat straw were 1.8-fold higher than that of the untreated wheat straw, under optimal enzymatic hydrolysis conditions. In addition, a maximum bioflocculant yield of 2.08 g L−1 was extracted from the fermentation broth after 15 days of incubation. The aforementioned bioflocculants could be used to efficiently decolorize a dye solution. Conclusions The results indicate that the cellulase-free xylanase-secreting P. boreopolis G22 may be a potential strain for wheat straw pretreatments. The strain G22 does not only enhance the enzymatic digestibility of wheat straw, but also simultaneously produces a number of bioflocculants that can be used for various industrial applications.

We have conducted a genome-level comparative study of basidiomycetes wood-rotting fungi (white, brown and soft rot) to understand the total plant biomass (lignin, cellulose, hemicellulose and pectin) -degrading abilities. We have retrieved the genome-level annotations of well-known 14 white rot fungi, 15 brown rot fungi and 13 soft rot fungi. Based on the previous literature and the annotations obtained from CAZy (carbohydrate-active enzyme) database, we have separated the genome-wide CAZymes of the selected fungi into lignin-, cellulose-, hemicellulose- and pectin-degrading enzymes. Results obtained in our study reveal that white rot fungi, especially Pleurotus eryngii and Pleurotus ostreatus potentially possess high ligninolytic ability, and soft rot fungi, especially Botryosphaeria dothidea and Fusarium oxysporum sp., potentially possess high cellulolytic, hemicellulolytic and pectinolytic abilities. The total number of genes encoding for cytochrome P450 monooxygenases and metabolic processes were high in soft and white rot fungi. We have tentatively calculated the overall lignocellulolytic abilities among the selected wood-rotting fungi which suggests that white rot fungi possess higher lignin and soft rot fungi potentially possess higher cellulolytic, hemicellulolytic and pectinolytic abilities. This approach can be applied industrially to efficiently find lignocellulolytic and aromatic compound-degrading fungi based on their genomic abilities.

Abstract Background Cellulases played an important role in the production of bioenergy and bio-products. Cellulases from bacteria with some special characteristics drew great attention due to its fast growth speed, wide adaption to harsh environment, and production of multi-function cellulases. Results An endoglucanase gene egls from Bacillus velezensis A4 was cloned and expressed in Escherichia coli BL21 (DE3). The recombinant enzyme Egls was partially purified using aqueous two-phase system. The highest recovery rate of the enzyme was 90.39% at PEG 4000 (25% w/w), phosphate buffer 8.08% (w/w) (pH 6.0), and NaCl (5% w/w). The enzyme molecular weight was 55 KD estimated by zymogram. The optimal pH and temperature of recombinant enzyme Egls were pH 6.0 and 55 °C, respectively. The enzyme was stable at pH range of 5.0–7.0 at 55 °C for 60 min. The enzyme exhibited K m, V max, K cat values as 63.38 mg/ml, 55.6 mg/min, and 3.93 × 103/S, respectively. The addition of 10 mM of Mg2+, Mn2+, or 5% (w/w) of Triton-X 100 in the reaction system enhanced the enzyme activity significantly. The enzyme showed both endoglucanase and exoglucanase activity. Conclusions An endoglucanase gene egls from B. velezensis A4 was cloned and expressed in E. coli BL21 (DE3). The recombinant enzyme Egls was purified by aqueous two-phase system and characterized. The enzyme can be applied for the efficient pretreatment of lignocellulosic biomass for bioenergy and bio-products production.

Abstract Background The thriving of biodiesel industry has led to produce 10% (v/v) crude glycerol, thus creating an overflow problem. Biofuel production is restricted by Escherichia coli due to its toxicity to bacterial cells. Therefore, a platform chemical and fuel additive 2,3-butanediol (2,3-BD) with low toxicity to microbes could be a promising alternative for biofuel production by recombinant E. coli using glycerol as the sole substrate. Results A novel expression system of E. coli was developed to express the dhaD gene encoding glycerol dehydrogenase (GDH) to produce value-added metabolic products through aerobic biotransformation of glycerol. The dhaD gene obtained from Klebsiella pneumoniae SRP2 was expressed in E. coli BL21(DE3)pLysS using an E. coli–K. pneumoniae shuttle vector pJET1.2/blunt consisting of chloramphenicol-resistance gene under the control of the T7lac promotor. RT-PCR analysis and dhaD overexpression confirmed that the 2,3-BD synthesis pathway gene was expressed on RNA and protein levels. Therefore, the recombinant E. coli exhibited a 38.9-fold higher enzyme activity (312.57 units/mg protein), yielding 8.97 g/L 2,3-BD, a 2.4-fold increase with respect to the non-recombinant strain. Conclusions The engineered strain E. coli BL21(DE3)pLysS/pJET1.2/blunt-dhaD, carrying the 2,3-BD pathway gene dhaD from our newly isolated Klebsiella pneumoniae SRP2 strain, displayed the best ability to synthesize 2,3-BD from low-cost biomass glycerol. The value of expression of an important glycerol metabolism gene dhaD is the highest ever achieved with an engineered E. coli strain. From these results, the first reported dhaD expression system has paved the way for improvement of 2,3-BD production and is efficient for another heterologous gene expression in E. coli.

Amylases account for about 30% of the global market of industrial enzymes, and the current amylases cannot fully meet industrial needs. This study aimed to identify a high α-amylase producing bacterium WangLB, to clone its α-amylase coding gene, and to characterize the α-amylase. Results showed that WangLB belonged to Bacillus velezensis whose α-amylase gene was 1980 bp coding 659 amino acids designated as BvAmylase. BvAmylase was a hydrophilic stable protein with a signal peptide and a theoretical pI of 5.49. The relative molecular weight of BvAmylase was 72.35 kDa, and was verified by SDS-PAGE. Its modeled structure displayed that it was a monomer composed of three domains. Its optimum temperature and pH were 70 °C and pH 6.0, respectively. It also showed high activity in a wide range of temperatures (40–75 °C) and a relatively narrow pH (5.0–7.0). It was a Ca2+-independent enzyme, whose α-amylase activity was increased by Co2+, Tween 20, and Triton X-100, and severely decreased by SDS. The Km and the Vmax of BvAmylase were 3.43 ± 0.53 and 434.19 ± 28.57 U/mg. In conclusion, the α-amylase producing bacterium WangLB was identified, and one of its α-amylases was characterized, which will be a candidate enzyme for industrial applications.

Abstract Background Bioflocculation has been developed as a cost-effective and environment-friendly method to harvest multiple microalgae. However, the high production cost of bioflocculants makes it difficult to scale up. In the current study, low-cost bioflocculants were produced from untreated corn stover by a biomass-degrading bacterium Pseudomonas sp. GO2. Results Pseudomonas sp. GO2 showed excellent production ability of bioflocculants through directly hydrolyzing various biomasses. The untreated corn stover was selected as carbon source for bioflocculants’ production due to its highest flocculating efficiency compared to that when using other biomasses as carbon source. The effects of fermentation parameters on bioflocculants’ production were optimized via response surface methodology. According to the optimal model, an ideal flocculating efficiency of 99.8% was obtained with the fermentation time of 130.46 h, initial pH of 7.46, and biomass content of 0.64%. The relative importance of carboxymethyl cellulase and xylanase accounted for 51.8% in the process of bioflocculants’ production by boosted regression tree analysis, further indicating that the bioflocculants were mainly from the hydrolysates of biomass. Biochemical analysis showed that it contained 59.0% polysaccharides with uronic acid (34.2%), 32.1% protein, and 6.1% nucleic acid in the bioflocculants, which had an average molecular weight as 1.33 × 106 Da. In addition, the bioflocculants showed the highest flocculating efficiency at a concentration of 12.5 mg L−1 and were stable over broad ranges of pH and temperature. The highest flocculating efficiencies obtained for Chlorella zofingiensis and Neochloris oleoabundans were 77.9 and 88.9%, respectively. Conclusions The results indicated that Pseudomonas sp. GO2 can directly utilize various untreated lignocellulolytic biomasses to produce low-cost bioflocculants, which showed the high efficiency to harvest two green microalgae in a low GO2 fermentation broth/algal culture ratio.

A xylanase producing bacterium was isolated from paper mill sludge in Thunder Bay, Canada. The newly isolated bacterium was identified as Pseudomonas boreopolis according to its 16S rRNA gene sequence and designated as LUQ1. The zymogram analysis indicated that there was one band of protein with xylanase activity, and the molecular weight of the enzyme was about 20 kDa. This xylanase works best at pH 6.0 and 65°C. The xylanase can be induced to express by xylose. The expression was enhanced on increasing the concentration of xylose, which reached its highest activity at 12 mM of xylose in fermentation. Wheat bran was the best carbon source in submerged fermentation. The highest xylanase activity of 25.61 U/ml was obtained at 96 h using wheat bran feedstock. When barley straw and wheat bran were used as feedstocks, the addition of 10 mM of xylose increased xylanase activity by ~50% and ~15%, respectively. The results showed that the strain LUQ1 has a great potential to produce xylanase for industrial applications.

Pectinases are a group of enzymes with broad application, including in plant fiber processing, pectic wastewater treatment, paper pulping, fruit juice extraction, and clarification. With an increasing industrial demand for these enzymes, it is useful to isolate organisms that produce large amounts of pectinase and possess wide ranges of stability factors like temperature and pH. In this study, 17 out of 29 bacteria (58.62%) from forest soil samples were pectinolytic. However, only four bacteria (S-5, S-10, S-14, and S-17) showed high pectin hydrolysis zones (ranging from 0.2 cm to 1.7 cm). These four bacteria were identified based on colony morphology, microscopic characterization, biochemical characteristics, and 16S rDNA sequencing. They were designated as Streptomyces sp. (S-5, S-14), Cellulomonas sp. (S-10), and Bacillus sp. (S-17). Interestingly, bacteria showed cellulase and xylanase activity in addition to pectinase. The quantitative assay for pectinase activity of the four isolates provided proof that they are pectinase producers and can be considered potential candidates for industrial uses. The crude enzyme extracts of these bacteria are applicable in oil and juice extraction from sesame seeds and apples, respectively.

As the condenser has complicated indoor environment and it’s difficult for the cleaning robot to complete cleaning, this paper deeply researches tele-operation system of intelligent cleaning robot, designs hardware architecture and software system at tele-operation control platform, including design of human-machine interface on the tele-operation control platform and software and hardware design of joystick force feedback controller. It designs the ontology mechanical structure, electromechanical control system, communication control program and picture processing program of intelligent cleaning robot. Industrial personal computer is used for data processing and motion control in the electromechanical control system. Test results show that the operation system herein can be used to effectively solve the problems in design and control of tele- operation system of intelligent cleaning robot and implement online high-pressure water-jetting cleaning of a condenser tube, so as to ensure efficient and safe operation of condensers in the large power plant.

In this study, crude lignin extracted from the black liquor generated by a pulp and paper mill was modified by different doses of irradiation. The crude and irradiation-modified lignins were used to treat wastewater that was generated during the production of starch glucoamylase. Changes to the physical and chemical properties and structure of the irradiation-modified lignins were determined using scanning electron microscopy, solubility analysis, elemental analysis, analysis of phenolic hydroxyl group, ultraviolet–visible spectroscopy, and Fourier transform infrared spectroscopy. Irradiation reduced the phenolic hydroxyl content in the lignin but increased its solubility by about 40%; analysis revealed that irradiation also destroyed the skeletal structure of the benzene ring in the lignin. After four minutes of settling, the total nitrogen (TN) and chemical oxygen demand (COD) in the wastewater reached 7.0 mg/L and 1573.1 mg/L, respectively. The settled solids content and protein recovery were 1.12 g/L and 98%, respectively. This study suggested that irradiation-modified lignin extracted from black liquor generated in the pulp and paper industry can be used to treat wastewater from the production of starch glucoamylase.