Elucidating the inhibiting mechanism of pseudo-lignin on the enzymatic digestibility of cellulose by surface plasmon resonance.

To explore the interaction mechanism of pseudo-lignin (PL) with cellulase and its influence on cellulose hydrolysis, different PLs were extracted from pretreated bamboo holocellulose (HC) using different organic solvents. Meanwhile, the real-time interaction of PL and cellulase was analyzed using surface plasmon resonance (SPR). The results showed that the extraction effect of the tetrahydrofuran and 1, 4-dioxane/water solution on PL was more effective than the ethanol/water solution. The inhibition of PL fraction obtained from HC by acid pretreatment with higher temperature showed less effect on Avicel's enzymatic hydrolysis. SPR analysis revealed that PL formed at higher pretreatment temperature had a lower dissociation rate after adsorption with cellulase. Besides, the binding affinity of PL (160 °C) to cellulase was much greater than that of PL obtained from 180 °C, indicating PL extracted at higher temperature treated biomass is more easily dissociated from cellulase after binding.

Revealing the mechanism of surfactant-promoted enzymatic hydrolysis of dilute acid pretreated bamboo.

To improve the enzymatic digestibility of dilute acid pretreated bamboo residue (DABR), surfactants including PEG 4000 and Tween 80 were added to prevent the non-productive adsorption between residual lignin and enzyme. At the optimal loadings (e.g., 0.2 and 0.3 g surfactant/g lignin), the enzymatic digestibility of DABR improved from 29.4% to 64.6% and 61.6% for PEG 4000 and Tween 80, respectively. Furthermore, the promoting mechanism of these surfactants on enzymatic hydrolysis was investigated by real-time surface plasmon resonance (SPR) and fluorescence spectroscopy. Results from SPR analysis showed that Tween 80 outperformed PEG 4000 in terms of dissociating the irreversible cellulase adsorption onto lignin. Fluorescence quenching mechanism revealed that PEG 4000 and Tween 80 intervened the interaction between lignin and cellulase by hydrogen bonds/Van der Waals and hydrophobic action, respectively. This work provided an in-depth understanding of the mechanisms of PEG 4000 and Tween 80 on enhancing the enzymatic hydrolysis efficiency.

Understanding the effects of different residual lignin fractions in acid-pretreated bamboo residues on its enzymatic digestibility.

BACKGROUND: During the dilute acid pretreatment process, the resulting pseudo-lignin and lignin droplets deposited on the surface of lignocellulose and inhibit the enzymatic digestibility of cellulose in lignocellulose. However, how these lignins interact with cellulase enzymes and then affect enzymatic hydrolysis is still unknown. In this work, different fractions of surface lignin (SL) obtained from dilute acid-pretreated bamboo residues (DAP-BR) were extracted by various organic reagents and the residual lignin in extracted DAP-BR was obtained by the milled wood lignin (MWL) method. All of the lignin fractions obtained from DAP-BR were used to investigate the mechanism for interaction between lignin and cellulase using surface plasmon resonance (SPR) technology to understand how they affect enzymatic hydrolysis RESULTS: The results showed that removing surface lignin significantly decreased the yield for enzymatic hydrolysis DAP-BR from 36.5% to 18.6%. The addition of MWL samples to Avicel inhibited its enzymatic hydrolysis, while different SL samples showed slight increases in enzymatic digestibility. Due to the higher molecular weight and hydrophobicity of MWL samples versus SL samples, a stronger affinity for MWL (KD = 6.8-24.7 nM) was found versus that of SL (KD = 39.4-52.6 nM) by SPR analysis. The affinity constants of all tested lignins exhibited good correlations (r > 0.6) with the effects on enzymatic digestibility of extracted DAP-BR and Avicel. CONCLUSIONS: This work revealed that the surface lignin on DAP-BR is necessary for maintaining enzyme digestibility levels, and its removal has a negative impact on substrate digestibility.

Enhancing the enzymatic digestibility of bamboo residues by biphasic phenoxyethanol-acid pretreatment.

The high content of lignin in bamboo is considered as the major obstacle for its biorefining. In this work, a green, lignin-selective, and recyclable solvent of phenoxyethanol was coupled with acid solution to deconstruct recalcitrant structure of bamboo residues (BR) to boost its enzymatic digestibility. Results showed phenoxyethanol has excellent lignin-removal ability from 29.4% to 91.6% when phenoxyethanol:acid ratios increased from 0:1 to 4:1 at 120 °C. 82.5%-87.8% of cellulose was preserved in pretreated BR. The enzymatic digestibility of BR significantly improved from 20.0% to 91.3% when it was pretreated under optimized conditions. With lower enzyme dosages (10 FPU/g) and 5 recycled using of pretreatment liquor, pretreated BR still showed a good enzymatic digestibility of 67.4%-93.7% and 67.1-76.8%, respectively. Examination of physicochemical changes revealed that improvements to accessibility, reduction of crystallite size, decrease of surface lignin and hydrophobicity for pretreated BR showed positive correlations (R2 > 0.7) with their enzymatic digestibility.

Removal of fermentation inhibitors from pre-hydrolysis liquor using polystyrene divinylbenzene resin.

BACKGROUND: The presence of soluble lignin, furfural and hydroxymethylfurfural (HMF) in industrial pre-hydrolysis liquor (PHL) from the pulping process can inhibit its bioconversion into bioethanol and other biochemicals. Although various technologies have been developed to remove these inhibitors, certain amounts of sugars are also inevitably removed during the treatment process. Hence, polystyrene divinylbenzene (PS-DVB) resin was used as an adsorptive material to simultaneously remove fermentation inhibitors while retaining sugars with high yields to improve the fermentability of PHL after acid hydrolysis by enriching its xylose concentration. The fermentability of acid-hydrolyzed PHL (A-PHL) was evaluated by the bioconversion into ethanol and xylosic acid (XA) after treatment with PS-DVB resin. RESULTS: The results showed that the highest xylose concentration (101.1 g/L) in PHL could be obtained by acid hydrolysis at 100 °C for 80 min with 4% acid, while the concentration of fermentation inhibitors (furfural, HMF and lignin) in PHL could also be significantly improved during the acid-hydrolysis process. After treatment with PS-DVB resin, not only were 97% of lignin, 92% of furfural, and 97% of HMF removed from A-PHL, but also 96% of xylose was retained for subsequent fermentation. With resin treatment, the fermentability of A-PHL could be improved by 162-282% for ethanol production from A-PHL containing 30-50 g/L xylose and by 18-828% for XA production from A-PHL containing 90-150 g/L xylose. CONCLUSIONS: These results confirmed that PS-DVB resin can remove inhibitors from PHL before producing value-added products by bioconversion. In addition, this work will ideally provide a concept for producing value-added chemicals from pre-hydrolysis liquor, which is regarded as the waste stream in the pulping process.

Chemerin Suppresses Breast Cancer Growth by Recruiting Immune Effector Cells Into the Tumor Microenvironment.

Infiltration of immune cells into the tumor microenvironment (TME) can regulate growth and survival of neoplastic cells, impacting tumorigenesis and tumor progression. Correlations between the number of effector immune cells present in a tumor and clinical outcomes in many human tumors, including breast, have been widely described. Current immunotherapies utilizing checkpoint inhibitors or co-stimulatory molecule agonists aim to activate effector immune cells. However, tumors often lack adequate effector cell numbers within the TME, resulting in suboptimal responses to these agents. Chemerin (RARRES2) is a leukocyte chemoattractant widely expressed in many tissues and is known to recruit innate leukocytes. CMKLR1 is a chemotactic cellular receptor for chemerin and is expressed on subsets of dendritic cells, NK cells, and macrophages. We have previously shown that chemerin acts as a tumor suppressive cytokine in mouse melanoma models by recruiting innate immune defenses into the TME. Chemerin/RARRES2 is down-regulated in many tumors, including breast, compared to normal tissue counterparts. Here, using a syngeneic orthotopic EMT6 breast carcinoma model, we show that forced overexpression of chemerin by tumor cells results in significant recruitment of NK cells and T cells within the TME. While chemerin secretion by EMT6 cells did not alter their phenotypic behavior in vitro, it did significantly suppress tumor growth in vivo. To define the cellular effectors required for this anti-tumor phenotype, we depleted NK cells or CD8+ T cells and found that either cell type is required for chemerin-dependent suppression of EMT6 tumor growth. Finally, we show significantly reduced levels of RARRES2 mRNA in human breast cancer samples compared to matched normal tissues. Thus, for the first time we have shown that increasing chemerin expression within the breast carcinoma TME can suppress growth by recruitment of NK and T cells, thereby supporting this approach as a promising immunotherapeutic strategy.

A Chimeric Antibody against ACKR3/CXCR7 in Combination with TMZ Activates Immune Responses and Extends Survival in Mouse GBM Models.

Glioblastoma (GBM) is the least treatable type of brain tumor, afflicting over 15,000 people per year in the United States. Patients have a median survival of 16 months, and over 95% die within 5 years. The chemokine receptor ACKR3 is selectively expressed on both GBM cells and tumor-associated blood vessels. High tumor expression of ACKR3 correlates with poor prognosis and potential treatment resistance, making it an attractive therapeutic target. We engineered a single chain FV-human FC-immunoglobulin G1 (IgG1) antibody, X7Ab, to target ACKR3 in human and mouse GBM cells. We used hydrodynamic gene transfer to overexpress the antibody, with efficacy in vivo. X7Ab kills GBM tumor cells and ACKR3-expressing vascular endothelial cells by engaging the cytotoxic activity of natural killer (NK) cells and complement and the phagocytic activity of macrophages. Combining X7Ab with TMZ allows the TMZ dosage to be lowered, without compromising therapeutic efficacy. Mice treated with X7Ab and in combination with TMZ showed significant tumor reduction by MRI and longer survival overall. Brain-tumor-infiltrating leukocyte analysis revealed that X7Ab enhances the activation of M1 macrophages to support anti-tumor immune response in vivo. Targeting ACKR3 with immunotherapeutic monoclonal antibodies (mAbs) in combination with standard of care therapies may prove effective in treating GBM.

Perspectives of biotechnological production of L-ribose and its purification.

L-ribose is a non-natural and expensive sugar that can be used as an important intermediate for the synthesis of L-nucleoside analogues, which are used as antiviral drugs. In contrast to chemical production, biotechnological methods can produce L-ribose from biomass under environmentally friendly conditions. In this mini-review, various strategies for synthesizing L-ribose by applying microorganisms and their enzymes are discussed, including microbial biotransformation and biocatalysis by engineering bacteria. Furthermore, subsequent isolation-and-purification techniques, as an integral step in the whole process, are accordingly described, containing the special introduction of a promising strategy of L-ribose separation. Particularly, further researches and outlook for the improvement of L-ribose preparation was solely stressed. Compared with each method, this mini-review provides a panorama of respective advantages and disadvantages existing in them.