A biosensor based on oriented immobilization of an engineered l-glutamate oxidase on a screen-printed microchip for detection of l-glutamate in fermentation processes.

This paper describes an amperometric biosensor utilizing an engineered l-glutamate oxidase for glutamate monitoring in microbial fermentation processes. We designed a general immobilization strategy that utilized a chitin-binding domain (ChBD-tag) as a biotether to further immobilize l-glutamate oxidase (GLOX) in an oriented manner on a screen-printed Prussian blue nanocube microchip (PB/SPC) with the biopolymer chitosan. The improved l-glutamate biosensor exhibited an enhanced sensitivity of 53.4 µA L mmol-1 cm-2 and a linear range from 25 µmol/L to 300 µmol/L with a detection limit of 9 µmol/L, and retained 95 % of its initial activity after two weeks of usage. In addition, the as-prepared biosensor was applied for real-time monitoring of food ingredient l-glutamate concentration during the fermentation process, which was in good agreement with that of high-performance liquid chromatography (HPLC). Above all, the l-glutamate biosensor prepared by this method had high analytical performance, and could fully realize real-time and high-efficiency monitoring in glutamate fermentation.

Quercetin Alleviates Lipopolysaccharide-Induced Inflammatory Response in Bovine Mammary Epithelial Cells by Suppressing TLR4/NF-κB Signaling Pathway.

Bovine mastitis is one of the most common clinical diseases in dairy cows, causing huge economic losses to the dairy industry. Quercetin is an important flavonoid existing in many food resources, which has attracted widespread attention as a potential anti-inflammatory and antioxidant. However, the molecular mechanism of quercetin on inflammatory responses and oxidative stress in bovine mammary epithelial cells (BMECs) induced by lipopolysaccharide (LPS) remains unknown. The objective of this study was to investigate the effects of quercetin on inflammation responses, oxidative stress, and barrier function of BMEC induced by LPS. Our results showed that BMEC viability was not affected by treatment with 50 and 100 µg/ml of quercetin and 1 µg/ml of LPS compared with control group. The results of oxidative stress indicators and related genes of barrier function indicated that 100 µg/ml of quercetin effectively protected the BMECs from damage of oxidative and barrier induced by 1 µg/ml of LPS. Moreover, the messenger RNA (mRNA) expressions of pro-inflammatory cytokines TNF-α, IL-1ß, IL-6, and chemokines CXCL2, CXCL5, CCL5, and CXCL8 were markedly decreased in the LPS-treated bovine retinal endothelial cells (BRECs) with 100 µg/ml of quercetin relatively to LPS alone. More importantly, the mRNA expressions of toll-like receptor 4 (TLR4), CD14, myeloid differential protein-2 (MD2), and myeloid differentiation primary response protein (MyD88) genes involved in TLR4 signal pathway were significantly attenuated by the addition of quercetin in LPS-treated BMEC, suggesting that quercetin can inhibit the TLR4 signal pathway. In addition, immunocytofluorescence showed that quercetin significantly inhibited the nuclear translocation of NF-κB p65 in BMEC induced by LPS. Therefore, the protective effects of quercetin on inflammatory responses in LPS-induced BMEC may be due to its ability to suppress the TLR4-mediated NF-κB signaling pathway. These findings suggest that quercetin can be used as an anti-inflammatory reagent to treat mastitis induced by exogenous or endogenous LPS release.

Techniques for enhancing the tolerance of industrial microbes to abiotic stresses: A review.

The diversity of stress responses and survival strategies evolved by microorganism enables them to survive and reproduce in a multitude of harsh environments, whereas the discovery of the underlying resistance genes or mechanisms laid the foundation for the directional enhancement of microbial tolerance to abiotic stresses encountered in industrial applications. Many biological techniques have been developed for improving the stress resistance of industrial microorganisms, which greatly benefited the bacteria on which industrial production is based. This review introduces the main techniques for enhancing the resistance of microorganisms to abiotic stresses, including evolutionary engineering, metabolic engineering, and process engineering, developed in recent years. In addition, we also discuss problems that are still present in this area and offer directions for future research.

Application of eukaryotic and prokaryotic laccases in biosensor and biofuel cells: recent advances and electrochemical aspects.

Laccases exhibit a wide range of applications, especially in the electrochemical field, where they are regarded as a potential biotic component. Laccase-based biosensors have immense practical applications in the food, environmental, and medical fields. The application of laccases as biocathodes in enzymatic biofuel cells has promising potential in the preparation of implantable equipment. Extensive studies have been directed towards the potential role of fungal laccases as biotic components of electrochemical equipment. In contrast, the potential of prokaryotic laccases in electrochemistry has been not fully understood. However, there has been recent and rapid progress in the discovery and characterization of new types of prokaryotic laccases. In this review, we have comprehensively discussed the application of different sources of laccases as a biocatalytic component in various fields of application. Further, we described the potential of different types of laccases in bioelectrochemical applications.

Characteristics and metabolic pathway of acetamiprid biodegradation by Fusarium sp. strain CS-3 isolated from soil.

An acetamiprid-degrading fungus was isolated from contaminated soil and identified as Fusarium sp. strain CS-3 based on physiological, biochemical, and molecular analyses. Strain CS-3 exploited 50 mg/L as the sole carbon source in liquid culture, removing 98% in 96 h. Strain CS-3 retained its acetamiprid degradation abilities over a wide range of pH (5.0-8.0) and temperature (20-42 °C). HPLC-MS analysis showed that N'-[(6-chloropyridin-3-yl)methyl]-N-methylacetamide, 2-chloro-5-hydroxymethylpyridine, and 6-chloronicotinic acid were identified as the most predominant metabolites, forming the basis for a newly described acetamiprid degradation pathway. Strain CS-3 efficiently degraded 99.6% of 50 mg/kg acetamiprid in soil, indicating potential for soil remediation.

Surface Display of Bacterial Laccase CotA on Escherichia coli Cells and its Application in Industrial Dye Decolorization.

Laccase CotA from Bacillus subtilis 168 was successfully displayed on the membrane of Escherichia coli cells using poly-γ-glutamate synthetase A protein (PgsA) from B. subtilis as an anchoring matrix. Further analyses demonstrated that the fusion protein PgsA/CotA efficiently translocates to the cell surface of E. coli with an enzymatic activity of 65 U/108 cells. Surface-displayed CotA was shown to possess improved enzymatic properties compared with those of the wild-type CotA, including higher thermal stability (above 90% activity at 70 °C and nearly 40% activity at 90 °C after 5-h incubation) and stronger inhibitor tolerance (approximately 80 and 65% activity when incubated with 200 and 400 mM NaCl, respectively). Furthermore, the whole-cell system was demonstrated to have high enzymatic activity against anthraquinone dye, Acid Blue 62, triphenylmethane dye, Malachite Green, and azo dye, Methyl Orange with the decolorization percentages of 91, 45, and 75%, after 5-h incubation, respectively.