Insights into cell robustness against lignocellulosic inhibitors and insoluble solids in bioethanol production processes

Increasing yeast robustness against lignocellulosic-derived inhibitors and insoluble solids in bioethanol production is essential for the transition to a bio-based economy. This work evaluates the effect exerted by insoluble solids on yeast tolerance to inhibitory compounds, which is crucial in high gravity processes. Adaptive laboratory evolution (ALE) was applied on a xylose-fermenting Saccharomyces cerevisiae strain to simultaneously increase the tolerance to lignocellulosic inhibitors and insoluble solids. The evolved strain gave rise to a fivefold increase in bioethanol yield in fermentation experiments with high concentration of inhibitors and 10% (w/v) of water insoluble solids. This strain also produced 5% (P > 0.01) more ethanol than the parental in simultaneous saccharification and fermentation of steam-exploded wheat straw, mainly due to an increased xylose consumption. In response to the stress conditions (solids and inhibitors) imposed in ALE, cells induced the expression of genes related to cell wall integrity (SRL1, CWP2, WSC2 and WSC4) and general stress response (e.g., CDC5, DUN1, CTT1, GRE1), simultaneously repressing genes related to protein synthesis and iron transport and homeostasis (e.g., FTR1, ARN1, FRE1), ultimately leading to the improved phenotype. These results contribute towards understanding molecular mechanisms that cells might use to convert lignocellulosic substrates effectively.

Ozonized biochar filtrate effects on the growth of Pseudomonas putida and cyanobacteria Synechococcus elongatus PCC 7942

Background Biochar ozonization was previously shown to dramatically increase its cation exchange capacity, thus improving its nutrient retention capacity. The potential soil application of ozonized biochar warrants the need for a toxicity study that investigates its effects on microorganisms. Results In the study presented here, we found that the filtrates collected from ozonized pine 400 biochar and ozonized rogue biochar did not have any inhibitory effects on the soil environmental bacteria Pseudomonas putida, even at high dissolved organic carbon (DOC) concentrations of 300 ppm. However, the growth of Synechococcus elongatus PCC 7942 was inhibited by the ozonized biochar filtrates at DOC concentrations greater than 75 ppm. Further tests showed the presence of some potential inhibitory compounds (terephthalic acid and p-toluic acid) in the filtrate of non-ozonized pine 400 biochar; these compounds were greatly reduced upon wet-ozonization of the biochar material. Nutrient detection tests also showed that dry-ozonization of rogue biochar enhanced the availability of nitrate and phosphate in its filtrate, a property that may be desirable for soil application. Conclusion Ozonized biochar substances can support soil environmental bacterium Pseudomonas putida growth, since ozonization detoxifies the potential inhibitory aromatic molecules. Graphical Abstract

Effects of Inhibitory Compounds Present in Lignocellulosic Biomass Hydrolysates on the Growth of Bacillus subtilis

This study evaluated the individual and combined effects of inhibitory compounds formed during pretreatment of lignocellulosic biomass on the growth of Bacillus subtilis. Ten inhibitory compounds commonly present in lignocellulosic hydrolysates were evaluated, which included sugar degradation products (furfural and 5-hydroxymethylfurfural), acetic acid, and seven phenolic compounds derived from lignin (benzoic acid, vanillin, vanillic acid, ferulic acid, p-coumaric acid, 4-hydroxybenzoic acid, and syringaldehyde). For the individual inhibitors, syringaldehyde showed the most toxic effect, completely inhibiting the strain growth at 0.1 g/L. In the sequence, assays using mixtures of the inhibitory compounds at a concentration of 12.5% of their IC50 value were performed to evaluate the combined effect of the inhibitors on the strain growth. These experiments were planned according to a Plackett–Burman experimental design. Statistical analysis of the results revealed that in a mixture, benzoic acid and furfural were the most potent inhibitors affecting the growth of B. subtilis. These results contribute to a better understanding of the individual and combined effects of inhibitory compounds present in biomass hydrolysates on the microbial performance of B. subtilis. Such knowledge is important to advance the development of sustainable biomanufacturing processes using this strain cultivated in complex media produced from lignocellulosic biomass, supporting the development of efficient bio-based processes using B. subtilis.

Establishment of Kluyveromyces marxianus as a Microbial Cell Factory for Lignocellulosic Processes: Production of High Value Furan Derivatives

The establishment of lignocellulosic biorefineries is dependent on microorganisms being able to cope with the stressful conditions resulting from the release of inhibitory compounds during biomass processing. The yeast Kluyveromyces marxianus has been explored as an alternative microbial factory due to its thermotolerance and ability to natively metabolize xylose. The lignocellulose-derived inhibitors furfural and 5-hydroxymethylfurfural (HMF) are considered promising building-block platforms that can be converted into a wide variety of high-value derivatives. Here, several K. marxianus strains, isolated from cocoa fermentation, were evaluated for xylose consumption and tolerance towards acetic acid, furfural, and HMF. The potential of this yeast to reduce furfural and HMF at high inhibitory loads was disclosed and characterized. Our results associated HMF reduction with NADPH while furfural-reducing activity was higher with NADH. In addition, furans’ inhibitory effect was higher when combined with xylose consumption. The furan derivatives produced by K. marxianus in different conditions were identified. Furthermore, one selected isolate was efficiently used as a whole-cell biocatalyst to convert furfural and HMF into their derivatives, furfuryl alcohol and 2,5-bis(hydroxymethyl)furan (BHMF), with high yields and productivities. These results validate K. marxianus as a promising microbial platform in lignocellulosic biorefineries.

Label-free characterisation of amyloids and alpha-Synuclein polymorphs by exploiting their intrinsic fluorescence property

Conventional in vitro aggregation assays often involve tagging with extrinsic fluorophores which can interfere with aggregation. We propose the use of intrinsic amyloid fluorescence lifetime represented by model-free phasor plots, as a label-free assay to characterise amyloid structure. Intrinsic amyloid fluorescence arises from structured packing of β-sheets in amyloids and is independent of aromatic-based fluorescence. We show that different amyloids (i.e., α-Synuclein (αS), β-Lactoglobulin and TasA) and different polymorphic populations of αS (induced by aggregation in salt-free and salt buffers mimicking the intra-/extracellular environments) can be differentiated by their unique fluorescence lifetimes. Moreover, we observe that disaggregation of pre-formed fibrils of αS and βLG leads to increased fluorescence lifetimes, distinct to those of their fibrillar counterpart. Our assay presents a medium-throughput method for rapid classification of amyloids and their polymorphs (the latter of which recent studies have shown lead to different disease pathology), and for testing small molecule inhibitory compounds.

Jab1 (COPS5) as an Emerging Prognostic, Diagnostic and Therapeutic Biomarker for Human Cancer

Jab1 (C-Jun activation domain-binding protein-1) has been reported to be critically involved in regulating apoptosis, cell proliferation, cell cycle, thereby affecting numerous pathways, DDR (DNA damage response) regulation, and genomic instability. Jab1 (CSN5) dysregulation has been positively associated with oncogenesis via activating oncogenes and deactivating various tumor suppressors. Jab1 overexpression has been reported in several tumor forms, illuminating its potent efficacy in cancer progression and metastasis. Jab1 has instigated prompt research interest in elucidating inhibitors of numerous oncoproteins and oncogenes for chemotherapeutics. Our review has presented strong evidence for presenting the significance of Jab1 overexpression in numerous carcinomas and its involvement in modulating various signaling pathways for cancer cell survival. This review may project a new way for utilizing Jab1 as a strong target for developing potent inhibitory compounds targeting Jab1 that could be further utilized in chemoprevention with limited side effects. Altogether this review further confirmed the crucial involvement of Jab1 in carcinogenesis and tumorigenesis, displaying the strong potential of Jab1 as one of the potent cancer biomarkers.