Characterization of an ssDNA ligase and its application in aptamer circularization.

Aptamers are widely used in various biomedical areas as novel molecular recognition elements, however, short single-stranded DNA (ssDNA) or RNA oligonucleotides are easily degraded by nucleases in biological fluids. This problem can be solved by circularizing aptamers with circular ligases. Herein, a moderately thermostable ssDNA ligase was expressed and purified. The purified ligase showed good circularization activity for different length substrates and much higher circularization efficiency than T4 RNA ligase 1. Biochemical characterization revealed that the enzyme showed optimal circularization activity at pH 7.5 and 50 ᵒC. Mn2+ and Mg2+ increased enzyme circularization activity, with Mn2+ having higher activity than Mg2+. The optimal concentrations of Mn2+ and ligase were 1.25-2.5 mM and 0.02 nM, respectively. The kinetic parameters Km, Vmax and Kcat of ssDNA ligase were 1.16 µM, 10.71 µM/min, and 10.7 min-1, respectively. The ssDNA ligase efficiency was nucleotide-dependent, and 5'-G and 3'-T were the most ligase-favored terminal nucleotides. In addition, the affinity and stability of the circular aptamer were determined. The affinity constant (KD) was 4.9 µM, and the stability increased compared to its linear form. Molecular docking results showed that the circular aptamer bound to the target via two hydrogen bonds. This study provides a simple and efficient aptamer circularization modification method for improving aptamer stability and expanding its applications.

Silica and Selenium Nanoparticles Attract or Repel Scale Insects by Altering Physicochemical Leaf Traits.

Although nanoparticles have gained attention as efficient alternatives to conventional agricultural chemicals, there is limited knowledge regarding their effects on herbivorous insect behavior and plant physicochemistry. Here, we investigated the effects of foliar applications of nano-silica (SiO2NPs) and nano-selenium (SeNPs), and bulk-size silica (SiO2) on the choice behavior of the arrowhead scale insect on mandarin orange plants. One leaf of a bifoliate pair was treated with one of the three chemicals, while the other was treated with water (control). The respective SiO2, SeO2, calcium (Ca), and carbon (C) content levels in the leaf epidermis and mesophyll were quantified using SEM-EDX (or SEM-EDS); leaf toughness and the arrowhead scale density and body size were measured. First-instar nymphs preferred silica-treated leaves and avoided SeNP-treated leaves. SiO2 content did not differ between control and SiO2NP-treated leaves, but was higher in bulk-size SiO2-treated leaves. The SiO2 level in the control leaves was higher in the SiO2NP treatment compared with that in the control leaves in the bulk-size SiO2 treatment. Silica-treated leaves increased in toughness, but SeNP-treated leaves did not; leaf toughness increased with mesophyllic SiO2 content. The insect density per leaf increased with leaf toughness, SiO2 content and, in the SiO2NP treatment, with epidermal C content. There was no correlation between SeO2 content and insect density. This study highlights the potential uses of SeNPs as an insect deterrent and of silica for enhancing leaf toughness and attracting scale insects.

[Computer-aided aptamers screening technologies: a review].

The computer information technology that has penetrated into every aspect of our lives, can not only assist the screening of drugs, but also simulate the effect of drugs. At present, computer-aided technologies have been used to screen aptamers, which play an important role in improving the screening efficiency and screening high affinity binding aptamers. This review summarized the screening methods of aptamers through computer-aided sequence evaluation, structural analysis and molecular docking.

Proteomic Studies of the Mechanism of Cytotoxicity, Induced by Palytoxin on HaCaT Cells.

Palytoxin (PLTX) is a polyether marine toxin isolated from sea anemones. It is one of the most toxic nonprotein substances, causing many people to be poisoned every year and to die in severe cases. Despite its known impact on Na+,K+-ATPase, much still remains unclear about PLTX's mechanism of action. Here, we tested different concentrations of PLTX on HaCaT cells and studied its distributions in cells, its impact on gene expression, and the associated pathways via proteomics combined with bioinformatics tools. We found that PLTX could cause ferroptosis in HaCaT cells, a new type of programmed cell death, by up-regulating the expression of VDAC3, ACSL4 and NCOA4, which lead to the occurrence of ferroptosis. PLTX also acts on the MAPK pathway, which is related to cell apoptosis, proliferation, division and differentiation. Different from its effect on ferroptosis, PLTX down-regulates the expression of ERK, and, as a result, the expressions of MAPK1, MAP2K1 and MAP2K2 are also lower, affecting cell proliferation. The genes from these two mechanisms showed interactions, but we did not find overlap genes between the two. Both ferroptosis and MAPK pathways can be used as anticancer targets, so PLTX may become an anticancer drug with appropriate modification.