Biodegradation of bisphenol A by a Pichia pastoris whole-cell biocatalyst with overexpression of laccase from Bacillus pumilus and investigation of its potential degradation pathways.

Bisphenol A (BPA), an endocrine disrupter with estrogen activity, can infiltrate animal and human bodies through the food chain. Enzymatic degradation of BPA holds promise as an environmentally friendly approach while it is limited due to lower stability and recycling challenges. In this study, laccase from Bacillus pumilus TCCC 11568 was expressed in Pichia pastoris (fLAC). The optimal catalytic conditions for fLAC were at pH 6.0 and 80 °C, with a half-life T1/2 of 120 min at 70 °C. fLAC achieved a 46 % degradation rate of BPA, and possible degradation pathways were proposed based on identified products and reported intermediates of BPA degradation. To improve its stability and degradation capacity, a whole-cell biocatalyst (WCB) was developed by displaying LAC (dLAC) on the surface of P. pastoris GS115. The functionally displayed LAC demonstrated enhanced thermostability and pH stability along with an improved BPA degradation ability, achieving a 91 % degradation rate. Additionally, dLAC maintained a degradation rate of over 50 % after the fourth successive cycles. This work provides a powerful catalyst for degrading BPA, which might decontaminate endocrine disruptor-contaminated water through nine possible pathways.

A Design Method on Durable Asphalt Pavement of Flexible Base on Anti-Rutting Performance and Its Application.

To solve the durability of flexible base asphalt pavement, especially its anti-rutting problem, a design method on durable asphalt pavement of flexible base on anti-rutting performance was put forward in the paper, based on many experiments and calculations. Firstly, a method that asphalt could be selected according to penetration and the anti-rutting factor of its base asphalt was found, which solved the problem of the asphalt selection of the flexible base asphalt mixture design. Meanwhile, a method of skeleton-density structure gradation design was proposed based on the fractal void ratio of coarse aggregate, fractal volume of fine aggregate in coarse aggregate, penetration, fractal dimension of gradation particle size, and rutting tests, which effectively solved in advance the rutting and fatigue performance of flexible base asphalt mixtures. Then, on the basis of the fatigue damage, a calculation method of fatigue life was suggested, which solved the problem that the fatigue damage of asphalt mixtures rarely considered the combined effects of creep damage and fatigue damage. In addition, a calculation method of rutting was formulated based on vehicle dynamic load and ANSYS 16.0 software. Lastly, the feasibility of the design method on durable asphalt pavement of flexible base on anti-rutting performance was verified combining with the real engineering of a supporting project and several numerical calculations and tests.

Structure-Guided Engineering of a Protease to Improve Its Activity under Cold Conditions.

Bacillus proteases commonly exhibit remarkably reduced activity under cold conditions. Herein, we employed a tailored combination of a loop engineering strategy and iterative saturation mutagenesis method to engineer two loops for substrate binding at the entrance of the substrate tunnel of a protease (bcPRO) from Bacillus clausii to improve its activity under cold conditions. The variant MT6 (G95P/A96D/S99W/S101T/P127S/S126T) exhibited an 18.3-fold greater catalytic efficiency than the wild-type (WT) variant at 10 °C. Molecular dynamics simulations and dynamic tunnel analysis indicated that the introduced mutations extended the substrate-binding pocket volume and facilitated extra interactions with the substrate, promoting catalysis through binding in a more favorable conformation. This study provides insights and strategies relevant to improving the activities of proteases and supplies a novel protease with enhanced activity under cold conditions for the food industry to maintain the initial flavor and color of food and reduce energy consumption.

Qiangguyin inhibited fat accumulation in OVX mice through the p38 MAPK signaling pathway to achieve anti-osteoporosis effects.

Postmenopausal osteoporosis (PMOP) is a common bone disease characterized by decreased bone density and increased bone fragility due to decreased estrogen levels. Qiangguyin (QGY) is transformed from the famous traditional Chinese medicine BuShen Invigorating Blood Decoction. In this study, we used QGY to treat PMOP. We observed that QGY significantly reduced fat accumulation in the chondro-osseous junction. However, its specific mechanism of action remains unclear. To determine the specific molecular mechanism of QGY, we explored the pharmacological mechanism by which QGY reduces fat accumulation in the chondro-osseous junction through network pharmacological analysis. The active components and targets related to PMOP and QGY were screened from different databases, forming a composition-target-disease network. Next, a comprehensive analysis platform including protein-protein interaction (PPI) network, Gene Ontology (GO) enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were established. The results revealed that QGY inhibits adipogenic differentiation by activating the mitogen-activated protein kinase (MAPK) signaling pathway, thus reducing the accumulation of fat in the chondro-osseous junction. For further verification. In vitro and in vivo experiments were carried out. Our data showed that QGY significantly reversed the high expression of fatty acid binding protein 4 (FABP4) and peroxisome proliferator-activated receptor γ (PPARγ). Further, QGY prevents fat accumulation by inhibiting the expression of p38. In summary, the results of this study suggested that QGY-induced phenotypic changes are related to the activation of the p38 MAPK signaling pathway.