Catalytic promiscuity of O-methyltransferases from Corydalis yanhusuo leading to the structural diversity of benzylisoquinoline alkaloids.

O-methyltransferases play essential roles in producing structural diversity and improving the biological properties of benzylisoquinoline alkaloids (BIAs) in plants. In this study, Corydalis yanhusuo, a plant used in traditional Chinese medicine due to the analgesic effects of its BIA-active compounds, was employed to analyze the catalytic characteristics of O-methyltransferases in the formation of BIA diversity. Seven genes encoding O-methyltransferases were cloned, and functionally characterized using seven potential BIA substrates. Specifically, an O-methyltransferase (CyOMT2) with highly efficient catalytic activity of both 4'- and 6-O-methylations of 1-BIAs was found. CyOMT6 was found to perform two sequential methylations at both 9- and 2-positions of the essential intermediate of tetrahydroprotoberberines, (S)-scoulerine. Two O-methyltransferases (CyOMT5 and CyOMT7) with wide substrate promiscuity were found, with the 2-position of tetrahydroprotoberberines as the preferential catalytic site for CyOMT5 (named scoulerine 2-O-methyltransferase) and the 6-position of 1-BIAs as the preferential site for CyOMT7. In addition, results of integrated phylogenetic molecular docking analysis and site-directed mutation suggested that residues at sites 172, 306, 313, and 314 in CyOMT5 are important for enzyme promiscuity related to O-methylations at the 6- and 7-positions of isoquinoline. Cys at site 253 in CyOMT2 was proved to promote the methylation activity of the 6-position and to expand substrate scopes. This work provides insight into O-methyltransferases in producing BIA diversity in C. yanhusuo and genetic elements for producing BIAs by metabolic engineering and synthetic biology.

An ultrasensitive CH3NH3PbBr3 quantum dots@SiO2-based electrochemiluminescence sensing platform using an organic electrolyte for aflatoxin B1 detection in corn oil.

Mycotoxins contamination, especially aflatoxin B1 (AFB1) in edible oils, is a health hazard. Therefore, AFB1 trace analysis methods are urgently needed. Electrochemiluminescence (ECL) is a popular sensing method because of its low background interference and high sensitivity. However, existing ECL assays for AFB1 detection are based on aqueous rather than oil systems. Herein, we report a CH3NH3PbBr3 quantum dots (MAPB QDs)@SiO2-based ECL sensor for AFB1 quantification in corn oil using an organic electrolyte. The luminophore loading and stability of the MAPB QDs@SiO2 particles were significantly improved compared to those of bulky MAPB materials, resulting in an enhanced ECL response. Further, exploiting molecular imprinting technology, an ECL sensor for AFB1 detection with an ultra-low detection limit of 8.5 fg/mL was prepared. The reliability of the sensor was confirmed by comparable recoveries of corn oil samples with those obtained by high-performance liquid chromatography, indicating its potential for food safety evaluation.

Anti-Lung Cancer Activity of the Curcumin Analog JZ534 In Vitro.

This study investigated the anticancer effect of the curcumin analog JZ534 on lung cancer cell lines H460, A549, H1975, and HCC827. The antiproliferation effect of JZ534 was measured through the methylthiazoletetrazolium assay, and cell colony formation was observed. Cell cycle and apoptosis were determined by flow cytometry, and the preliminary mechanism was studied by Western blot. Results showed that JZ534 significantly inhibited the vitality and colony formation of lung cancer cells. JZ534 induced the G2/M cell cycle arrest of the cancer cells and suppressed the expression of cycle-related proteins, including cyclin B1 and Cdc2. Meanwhile, JZ534 induced cell apoptosis and upregulated the expression of apoptosis-related proteins, including cleaved caspase-3, Bax, and p53. At the same dose, JZ534 showed better antitumor activity than curcumin. These results suggest that JZ534 exhibits excellent anti-lung cancer activity by inhibiting the growth and inducing the apoptosis of lung cancer cells. Therefore, JZ534 is a promising lead compound for cancer treatment.

[Encircling needling combined with physical factor therapy for severe pressure sore].

OBJECTIVE: To compare the clinical efficacy difference between encircling needling combined with physical factor therapy and simple physical factor therapy for severe pressure sore, and to explore the optimal method for severe pressure sores. METHODS: Thirty-four patients with IV-grade pressure sore were randomly divided into an observation group and a control group, 17 cases in each one. Patients in the control group were treated with conventional nursing, ultrasonic wave and short-wave ultraviolet therapy; additionally, the encircling needling was applied in the observation group. All the treatment was given once a day, 5 times a week, and 4-week treatment constituted one session. Totally, two sessions of treatment were performed. Three indices, including the area of pressure sore, 24-h volume of exudates and wound-bed tissue type, were compared between the two groups before and after treatment; the clinical efficacy was evaluated in the two groups. RESULTS: After treatment of one session and two sessions, the area of pressure sore, 24-h volume of exudates and wound-bed tissue type were significantly reduced in the two groups (P < 0.01, P < 0.05), which was more obvious in the observation group (all P < 0.05). The total effective rate in the observation group was 76.5% (13/17) after 1 session and 94.1% (16/17) after 2 sessions, which were superior to 35.3% (6/17) after 1 session and 64.7% (11/17) after 2 sessions in the control group (both P < 0.05). CONCLUSION: Encircling needling combined with physical factor therapy can obviously reduce the pressure sore area and 24-h volume of exudates and improve wound-bed tissue type, which is superior to simple physical factor therapy.

Establishment of the active catalytic domain of human PDGFRbeta tyrosine kinase-based ELISA assay for inhibitor screening.

Tyrosine kinases are emerging as frequent targets of primary oncogenic events and therefore represent an optimal focus of therapeutic intervention. In an effort towards therapeutic PDGFR inactivation, we expressed the catalytic domain of PDGFRbeta as a soluble active kinase using Bac-to-Bac expression system, and studied the correlations between PDGFRbeta activity and enzyme concentration, ATP concentration, substrate concentration and divalent cation type. And a convenient, effective and non-radioactive ELISA screening model is then established for identification of the potential inhibitors targeting PDGFRbeta kinase. Of 500 RTK target-based compounds, TKI-30 was identified as a small molecule potential inhibitor of PDGFRbeta (IC(50)=0.34 microM). Further studies indicated that TKI-30 blocked PDGF-BB-induced autophosphorylation of PDGFRbeta in a dose-dependent manner in Swiss 3T3 cells and human umbilical vein smooth muscle cells (HUVSMCs). Moreover, it dose-dependently suppressed the PDGF-BB-induced proliferation in HUVSMCs and tube formation of HUVEC. Our data collectively indicated that PDGFRbeta-based ELISA assay is a new method available for screening inhibitors targeting PDGFRbeta kinase and TKI-30 is a potential novel anti-cancer agent worthy of being further investigated.

Evaluation of active recombinant catalytic domain of human ErbB-2 tyrosine kinase, and suppression of activity by a naturally derived inhibitor, ZH-4B.

Human cancers frequently express high levels of ErbB-2 tyrosine kinase, which is associated with aggressive tumor behavior and poor prognosis. ErbB-2 is thus a promising target for cancer therapy. Here we express the catalytic domain of ErbB-2 as a soluble active kinase, and investigate the correlations between its activity and kinase concentration, ATP concentration, substrate concentration and divalent cation type. A simple and effective screening model is established to identify and evaluate potential inhibitors of ErbB-2 kinase. ZH-4B, a naturally derived small molecule compound that potently inhibits ErbB-2 kinase activity with an IC50 value of 2.45+/-0.56 microM, is identified. In SK-OV-3 human ovarian cancer cells and SK-BR-3 human breast carcinoma cells, ZH-4B blocks epidermal growth factor (EGF)-induced phosphorylation of ErbB-2 in a dose-dependent manner. Our data collectively indicate that ZH-4B is a potential novel anti-cancer agent that deserves further investigation.