Ubiquitin-specific protease 15 (USP15) regulates important cellular processes, including transforming growth factor β (TGF-β) signaling, mitophagy, mRNA processing, and innate immune responses; however, structural information on USP15's catalytic domain is currently unavailable. Here, we determined crystal structures of the USP15 catalytic core domain, revealing a canonical USP fold, including a finger, palm, and thumb region. Unlike for the structure of paralog USP4, the catalytic triad is in an inactive configuration with the catalytic cysteine ∼10 Å apart from the catalytic histidine. This conformation is atypical, and a similar misaligned catalytic triad has so far been observed only for USP7, although USP15 and USP7 are differently regulated. Moreover, we found that the active-site loops are flexible, resulting in a largely open ubiquitin tail–binding channel. Comparison of the USP15 and USP4 structures points to a possible activation mechanism. Sequence differences between these two USPs mainly map to the S1′ region likely to confer specificity, whereas the S1 ubiquitin–binding pocket is highly conserved. Isothermal titration calorimetry monoubiquitin- and linear diubiquitin-binding experiments showed significant differences in their thermodynamic profiles, with USP15 displaying a lower affinity for monoubiquitin than USP4. Moreover, we report that USP15 is weakly inhibited by the antineoplastic agent mitoxantrone in vitro. A USP15–mitoxantrone complex structure disclosed that the anthracenedione interacts with the S1′ binding site. Our results reveal first insights into USP15's catalytic domain structure, conformational changes, differences between paralogs, and small-molecule interactions and establish a framework for cellular probe and inhibitor development.
Previous research has shown that some compounds in leaves and seeds of sugar apple have a cytotoxic activity. The aim of this research was to determine the cytotoxicity of semipolar fraction of ethanolic extract from sugar apple stem bark (Annona squamosa L.) on T47D cancer cells. The semipolar fraction of ethanolic extract from sugar apple stem bark was collected by fractionation using Vacuum Liquid Chromatography (VLC) with hexane:ethyl acetic (9:1, 8:2, 7:3, and 6:4) as mobile phase. Cytotoxicity from the fractions of five different concentration namely; 25, 50, 100, 150, and 250, µg/mL was measured by MTT assay. The potency of the cytotoxicity was defined by the ability of the fraction to inhibit the growth of T47D cells indicated by the value of IC50. Qualitative analysis of contained compounds in the fraction was done by Thin Layer Chromatography (TLC) method using silica gel F 254 as a stationary phase and hexane:ethyl acetic (7:3) as a mobile phase. UV 254 and 366 nm lamp also Dragendorff, citroboric, and FeCl3 spray reagents were used to visualize the spots of the secondary metabolites. The result proved that the semipolar fraction of ethanolic extract from sugar apple stem bark showed potential cytotoxicity on T47D cancer cells with IC50value of 70,77 µg/mL. Qualitative analysis showed that the fraction contained flavonoids and alkaloids which is presumably responsible for its cytotoxic activity.
Cynometra ramiflora Linn traditionally used as anti diabetic, anti hyperuricemia, hypertension, rheumatoid, and others diseases. It is important to obtain other biological activity of Sala plant. Previous research found that the ethanol extract of the steam bark Sala plant from Bangladesh has antibacterial activity. The aim of this study is to evaluate the antibacterial activity of ethanolic extract of steam bark of Sala plant against five bacterial species based on value of MBC. Antibacterial evaluated by solid dilution method using Mueller Hinton (MH) media. The test divided to media control, solvent and bacteria suspension and the extract group. The extract screened against five bacteria (Shigella sonei, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli multi resistant and Staphylococcus aureus multi resistant) at four different concentration (0.125; 0.25, 1 and 2%). The results showed that the ethanol extract of steam bark of Cynometra ramiflora Linn has potential as an antibacterial activity with MBC value of 2% against S. aureus, P. aeruginosa and S. sonei. On the other hand, K pneumonia and E. coli exhibited low potential because until highest concentration.
Four compounds, namely β-sitosterol (1), betulinic acid (2), cinnamic acid (3), and α-viniferin (4) have been successfully isolated from the bark methanol extract of Dipterocarpus confertus Sloot. The structures of the isolated compounds have been established on the basis spectroscopic data evidence, and comparison with the published data. In the cytotoxicity study, cinnamic acid (3) and betulinic acid (2) have been found to be very strong active against murine leukemia P388 and vero cells lines with the IC50 values of 2.25 and 5.10 µg/mL, respectively, while the other compounds were not active.
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