Molecular Docking and Site-directed Mutagenesis of a Bacillus thuringiensis Chitinase to Improve Chitinolytic,Synergistic Lepidopteran-larvicidal and Nematicidal Activities

Bacterial chitinases are useful in the biocontrol of agriculturally important pests and fungal pathogens. However, the utility of naturally occurring bacterial chitinases is often limited by their low enzyme activity. In this study, we constructed mutants of a Bacillus thuringiensis chitinase with enhanced activity based on homology modeling, molecular docking, and the site-directed mutagenesis of target residues to modify spatial positions, steric hindrances, or hydrophilicity/hydrophobicity. We first identified a gene from B. thuringiensis YBT-9602 that encodes a chitinase (Chi9602) belonging to glycosyl hydrolase family 18 with conserved substrate-binding and substrate-catalytic motifs. We constructed a structural model of a truncated version of Chi9602 (Chi9602_35-459) containing the substrate-binding domain using the homologous 1ITX protein of Bacillus circulans as the template. We performed molecular docking analysis of Chi9602_35-459 using di-N-acetyl-D-glucosamine as the ligand. We then selected 10 residues of interest from the docking area for the site-directed mutagenesis experiments and expression in Escherichia coli. Assays of the chitinolytic activity of the purified chitinases revealed that the three mutants exhibited increased chitinolytic activity. The ChiW50A mutant exhibited a greater than 60 % increase in chitinolytic activity, with similar pH, temperature and metal ion requirements, compared to wild-type Chi9602. Furthermore, ChiW50A exhibited pest-controlling activity and antifungal activity. Remarkable synergistic effects of this mutant with B. thuringiensis spore-crystal preparations against Helicoverpa armigera and Caenorhabditis elegans larvae and obvious activity against several plant-pathogenic fungi were observed.     

DFT comparison of the OH-initiated degradation mechanisms for five chlorophenoxy herbicides

To compare the OH-initiated reaction mechanisms of five chlorophenoxy herbicides, density functional theory (DFT) calculations of reactions in which ·OH attacks one of three active positions on each herbicide were carried out at the MPWB1K/6-311 + G(3df,2p)//MPWB1K/6-31 + G(d,p) level. For each herbicide, the calculation results show that ·OH addition to the C1 atom, which is the nexus between the benzene ring and the side group, possesses the lowest energy barrier among the three kinds of reactions, indicating that ·OH addition–substitution of the side chain is the most energetically and kinetically favorable reaction mechanism. Comparisons among the herbicides show that the mechanisms are affected by the steric hindrance and the electronegativities of the –CH₃ and –Cl groups. When comparing the addition of ·OH to the C1 site among the five herbicides, the activation energy for the reaction of ·OH with DCPP reaction is the lowest (3.61 kcal mol^?1), while that for the ·OH and 4-CPA reaction was the highest (5.91 kcal mol^?1). ·OH addition to the C4 site presents the highest energy barriers among the three kinds of reactions, indicating that the para Cl is difficult to break down. When comparing the H-atom abstraction reactions of the five herbicides, the H atoms in the –CH₂– group of 2,4-D are the easiest for ·OH to abstract, whereas those of DCPP and MCPP are more difficult to abstract, due to the steric hindrance of the –CH₃ group. Additionally, the results obtained from the PCM calculations reveal that most of the reactions occur more easily in water than in gas, though the mechanisms involved are the same as those discussed above.     

Looking into the grooves of DNA

The present study is aimed at understanding the effects of DNA sequence, local conformation, and curvature on groove geometry. Energy-optimized structures are obtained by Jumna methodology; groove geometry is analyzed by a recently developed technique that allows an accurate and continuous measurement of width and depth. The mechanics of groove deformations is also studied and analyzed in terms of helicoidal parameters. © 1993 John Wiley & Sons, Inc.     

Amino acids-Rab1A-mTORC1 signaling controls whole-body glucose homeostasis

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A (1.5 + ε)-Approximation Algorithm for Unsigned Translocation Distance

Genome rearrangement is an important area in computational biology and bioinformatics. The translocation operation is one of the popular operations for genome rearrangement. It was proved that computing the unsigned translocation distance is NP-hard. In this paper, we present a (1.5+epsiv)-approximation algorithm for computing the unsigned translocation distance that improves upon the best known 1.75-ratio. The runtime of our algorithm is O(n²+(4/epsiv)^1.5radic(log(4/epsiv)2^4/epsiv)), where n is the total number of genes in the genome.     

Effect of static magnetic field on morphology and growth metabolism of Flavobacterium sp. m1-14

Increasing evidence shows that static magnetic fields (SMFs) can affect microbial growth metabolism, but the specific mechanism is still unclear. In this study, we have investigated the effect of moderate-strength SMFs on growth and vitamin K₂ biosynthesis of Flavobacterium sp. m1-14. First, we designed a series of different moderate-strength magnetic field intensities (0, 50, 100, 150, 190 mT) and exposure times (0, 24, 48, 72, 120 h). With the optimization of static magnetic field intensity and exposure time, biomass and vitamin K₂ production significantly increased compared to control. The maximum vitamin K₂ concentration and biomass were achieved when exposed to 100 mT SMF for 48 h; compared with the control group, they increased by 71.3% and 86.8%, respectively. Interestingly, it was found that both the cell viability and morphology changed significantly after SMF treatment. Second, the adenosine triphosphate (ATP) and glucose-6-phosphate dehydrogenase (G6PDH) metabolism is more vigorous after exposed to 100 mT SMF. This change affects the cell energy metabolism and fermentation behavior, and may partially explain the changes in bacterial biomass and vitamin K₂ production. The results show that moderate-strength SMFs may be a promising method to promote bacterial growth and secondary metabolite synthesis.

     

生物类专业基础课“生物化学”课程思政教育探索与创新

专业课教学中渗透思政教育是实现高校全过程、全方位、全员育人的重要途径，而且以专业知识为载体的思政教育比纯粹的思政课更有说服力和感染力。生物化学作为生物类专业的基础必修课，蕴含着丰富多样的思政资源。近年来，我们教学团队结合“生物化学”课程自身特征，开展了颇有成效的课程思政教学探索与实践，明确了课程育人目标，丰富了课程教学资源，创新了课程教学模式，构建了思政融合策略。思政资源的深度挖掘和巧妙应用收获了“润物细无声”的效果，实现了“生物化学”课程教学价值塑造、知识传授与能力培养的三者有机结合。