ABSTRACT
Novel N-chloro â-Lactam and benzisoxazole derivatives were successfully synthesized with excellent yields (92-96%) under simple and mild reaction conditions. The ß-lactams as a class acquired importance since the discovery of penicillin which contains ß-lactam unit as an essential structural feature of its molecule, this interest continued unabated because of the therapeutic importance of ß-lactam antibiotics. In silico studies of the compounds with cancer drug target enzymes showed the inhibition of HDAC (Histone Deacetylase) and NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) significantly. The compounds were then investigated for the inhibitory potential against the same enzymes in vitro. NF-κB inhibition was investigated by trans activation assay using HEK293/NF-κB-luc cells. Overall, the synthesized compounds induce the cancer cell toxicity by restraining the NF-κB transcription factor mediated by HDAC inhibition and thus the compounds act as dual inhibitors.
Subject(s)
Histone Deacetylases/metabolism , Isoxazoles/chemistry , Isoxazoles/pharmacology , NF-kappa B/metabolism , beta-Lactams/chemistry , beta-Lactams/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Antineoplastic Agents/metabolism , Antineoplastic Agents/pharmacology , Catalysis , Catalytic Domain , HEK293 Cells , Hemolysis/drug effects , Histone Deacetylase Inhibitors/chemical synthesis , Histone Deacetylase Inhibitors/chemistry , Histone Deacetylase Inhibitors/metabolism , Histone Deacetylase Inhibitors/pharmacology , Histone Deacetylases/chemistry , Humans , Isoxazoles/chemical synthesis , Isoxazoles/metabolism , Models, Molecular , Molecular Docking Simulation , Stereoisomerism , beta-Lactams/chemical synthesis , beta-Lactams/metabolismABSTRACT
In the course of systematic screening program for bioactive actinomycetes, an alkaline protease producing halophilic strain Actinopolyspora sp. VITSDK2 was isolated from marine saltern, Southern India. The strain was identified as Actinopolyspora based on its phenotypic and phylogenetic characters. The protease was partially purified using ammonium sulfate precipitation and subsequently by DEAE cellulose column chromatography. The enzyme was further purified using HPLC and the molecular weight was found to be 22 kDa as determined by SDS-PAGE analysis. The purified protease exhibited pH stability in a wide range of 4-12 with optimum at 10.0. The enzyme was found to be stable between 25 and 80 °C and displayed a maximum activity at 60 °C. The enzyme activity was increased marginally in presence of Mn(2+) , Mg(2+) , and Ca(2+) and decreased in presence of Cu(2+) . PMSF and DFP completely inhibited the activity suggesting it belongs to serine protease. Further, the proteolytic activity was abolished in presence of N-tosyl-L-lysine chloromethyl ketone suggesting this might be chymotrypsin-like serine protease. The protease was 96% active when kept for 10 days at room temperature. The results indicate that the enzyme belong to chymotrypsin-like serine protease exhibiting both pH and thermostability, which can be used for various applications in industries.
Subject(s)
Actinobacteria/enzymology , Actinobacteria/isolation & purification , Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Endopeptidases/chemistry , Endopeptidases/metabolism , Geologic Sediments/microbiology , Actinobacteria/classification , Actinobacteria/metabolism , Bacterial Proteins/isolation & purification , Endopeptidases/isolation & purification , Enzyme Stability , India , Salinity , Serine Proteases/chemistry , Serine Proteases/isolation & purification , Serine Proteases/metabolismABSTRACT
The present study was designed to investigate the Puducherry coast of the Bay of Bengal, India for the diversity of bioactive actinomycetes. A total of 50 actinomycete strains were isolated from the marine sediments and most of the strains were belongs to Streptomyces. These strains were identified by means of morphological physiological, biochemical and cultural characteristics. The isolates were subjected to shake flask fermentation and the secondary metabolites were extracted with ethyl acetate and screened for cytotoxicity, hemolytic activity and antimicrobial activity against selected bacterial and fungal pathogens. The cytotoxic activity was evaluated using HeLa cell lines by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a tetrazole (MTT) assay, hemolytic activity on mouse erythrocytes and the antifungal activity was evaluated by MTT cytotoxic assay against Aspergillus niger, Aspergillus fumigatus and Candida albicans. The antibacterial activity was studied against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Klebsiella pneumoniae. The cytotoxicity and antimicrobial activity of secondary metabolite was found to be concentration dependent and nearly 24% of isolates showed significant antimicrobial, hemolytic and cytotoxic activity. The results of our study indicate the diversity and bioactive potential of marine actinomycetes isolated in the Puducherry coast.
