Site-directed mutagenesis improves the practical application of L-glutamic acid decarboxylase in Escherichia coli

γ-Aminobutyric acid (GABA) is a kind of non-proteinogenic amino acid which is highly soluble in water and widely used in the food and pharmaceutical industries. Enzymatic conversion is an efficient method to produce GABA, whereby glutamic acid decarboxylase (GAD) is the key enzyme that catalyzes the process. The activity of wild-type GAD is usually limited by temperature, pH or biotin concentration, and hence directional modification is applied to improve its catalytic properties and practical application. GABA was produced using whole cell transformation of the recombinant strains Escherichia coli BL21(DE3)-Gad B, E. coli BL21(DE3)-Gad B-T62S and E. coli BL21(DE3)-Gad B-Q309A. The corresponding GABA concentrations in the fermentation broth were 219.09, 238.42, and 276.66 g/L, and the transformation rates were 78.02%, 85.04%, and 98.58%, respectively. The results showed that Gad B-T62S and Gad B-Q309A are two effective mutation sites. These findings may contribute to ideas for constructing potent recombinant strains for GABA production. Practical Application : Enzymatic properties of the GAD from Escherichia coli and GAD site-specific mutants were examined by analyzing their conserved sequences, substrate contacts, contact between GAD amino acid residues and mutation energy (ΔΔG) of the GAD mutants. The enzyme activity and stability of Gad B-T62S and Gad B-Q309A mutants were improved compared to Gad B. The kinetic parameters K_m and V_max of Gad B, Gad B-T62S, and Gad B-Q309A mutants were 11.3 ± 2.1 mM and 32.1 ± 2.4 U/mg, 7.3 ± 2.5 mM and 76.1 ± 3.1 U/mg, and 7.2 ± 3.8 mM and 87.3 ± 1.1 U/mg, respectively. GABA was produced using whole cell transformation of the recombinant strains E. coli BL21(DE3)-Gad B, E. coli BL21(DE3)-Gad B-T62S, and E. coli BL21(DE3)-Gad B-Q309A. The corresponding GABA concentrations in the fermentation broth were 219.09, 238.42, and 276.66 g/L, and the transformation rates were 78.02%, 85.04%, and 98.58%, respectively.     

Assessing the suitability for Aedes albopictus and dengue transmission risk in China with a delay differential equation model

Dengue is considered non-endemic to mainland China. However, travellers frequently import the virus from overseas and local mosquito species can then spread the disease in the population. As a consequence, mainland China still experiences large dengue outbreaks. Temperature plays a key role in these outbreaks: it affects the development and survival of the vector and the replication rate of the virus. To better understand its implication in the transmission risk of dengue, we developed a delay differential equation model that explicitly simulates temperature-dependent development periods and tested it with collected field data for the Asian tiger mosquito, Aedes albopictus. The model predicts mosquito occurrence locations with a high accuracy (Cohen’s κ of 0.78) and realistically replicates mosquito population dynamics. Analysing the infection dynamics during the 2014 dengue outbreak that occurred in Guangzhou showed that the outbreak could have lasted for another four weeks if mosquito control interventions had not been undertaken. Finally, we analyse the dengue transmission risk in mainland China. We find that southern China, including Guangzhou, can have more than seven months of dengue transmission per year while even Beijing, in the temperate north, can have dengue transmission during hot summer months. The results demonstrate the importance of using detailed vector and infection ecology, especially when vector-borne disease transmission risk is modelled over a broad range of climatic zones.     

Synthesis and Anti-Oomycete Activity of 1-Sulfonyloxy/Acyloxydihydroeugenol Derivatives

Endeavor to discover biorational natural products-based fungicides, two series (26) of novel 1-sulfonyloxy/acyloxydihydroeugenol derivatives ( 3a – p and 5a – j ) were prepared and assessed for their fungicidal activity against P. capsici Leonian, in vitro. Results of fungicidal activity revealed that, among all compounds, especially compounds 3a , 5c , and 5e displayed the most potent anti-oomycete activity against P. capsici with EC₅₀ values of 69.33, 68.81, and 67.77 mg/L, respectively. Overall, the anti-oomycete activities of 1-acyloxydihydroeugenol derivatives ( 5a – j ) were higher than that of 1-sulfonyloxydihydroeugenol derivatives ( 3a – p ). It is proved that the introduction of the acyl group at hydroxy position of dihydroeugenol is more beneficial to improve its anti-oomycete activity than that of the sulfonyl group. These preliminary results will pave the way for further modification of dihydroeugenol in the development of potential new fungicides.     

The chromatin-remodeling protein BAF60/SWP73A regulates the plant immune receptor NLRs

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Biocompatible elastomer of waterborne polyurethane based on castor oil and polyethylene glycol with cellulose nanocrystals

Biocompatible waterborne polyurethane (WPU) based on castor oil (CO)/polyethylene glycol (PEG) filled with low level loadings of Eucalyptus globulus cellulose nanocrystals (ECNs) was prepared. The ECNs obtained by sulfuric hydrolysis, consisted of ‘rod-like’ crystals with an average length and diameter of 518.0 ± 183.4 nm and 21.7 ± 13.0 nm, respectively. The nanocomposites with low level loadings of ECNs showed significant enhancement in tensile strength and Young's modulus from 5.43 to 12.22 MPa and from 1.16 to 4.83 MPa, respectively. SEM results showed well dispersion of ECNs in the WPU matrix. Furthermore, it was verified that the nanosized ECNs favored the hard-segments (HSs)/soft-segments (SSs) microphase separation of the WPU, causing shifts of the SS glass transition temperature (T_g,s) and the HS melting temperature (T_m,h) toward higher temperatures.     

靶向递送蛋白水解靶向嵌合体在癌症治疗中的研究进展

小分子抗癌药物通过靶向特定蛋白来抑制肿瘤生长,但大部分致病蛋白被认为是“不可成药”的。蛋白水解靶向嵌合体(proteolysis targeting chimeras,PROTAC)通过靶向降解目标蛋白来抑制肿瘤细胞生长,是一项非常有潜力的新技术。本文在介绍传统多肽型PROTAC和小分子型PROTAC基础上,详细总结了靶向递送型PROTAC的最新研究进展,主要包括识别分子介导靶向PROTAC、纳米材料介导靶向PROTAC和可控激活小分子PROTAC前药。研究表明,靶向递送型PROTAC在提高肿瘤细胞特异性、减少脱靶效应和降低生物毒性等方面具有潜在应用价值。最后,本文对PROTAC的成药性进行了展望。