Project description:The crude enzyme extract from Artemisia annua was subjected to catalytic activity-guided purification and fractionation, yielding the active fraction DEAE-2, which demonstrated the biological function of catalyzing the interconversion between artemisinic acid and dihydroartemisinic acid. Proteomic identification of DEAE-2 was performed to uncover the functional enzymes responsible for this activity.
Project description:DNase III, also known as TREX1, is a 314 amino acid endoplasmic reticulum (ER) tail-anchored 3’ exonuclease where the N-terminal region contains the DNase domain and the C-terminal end controls TREX1 localization to the surface of the ER. Disease mutations in the C-terminal region alters TREX1 ability to interact with the oligosaccharyltransferase (OST) subunits Ribophorin 1 (RPN1) and DDOST leading to rapid hydrolysis of lipid-linked oligosaccharides (LLOs) into bioactive free oligosaccharides (fOS) in a switch-like manner. Bioassay-guided fractionation of the fOS pool revealed that the structure responsible for the bioactivity is a mannose (Man) b1-4 N-acetylglucosamine (GlcNAc) disaccharide. The bioactive disaccharide is produced from OST’s hydrolyzed LLOs in the cytoplasm and activates a STING-TBK1 dependent immune signal that leads to the upregulation of interferon-stimulated genes (ISGs) and chemokine genes.
Project description:HRMS guided study of 42 marine Streptomyces in the MAR4 clade. Metabolomics data collected from Ethyl Acetate crude extracts of 50 ml liquid cultures.
Project description:Top-down and bottom-up protein analysis of venom and saliva of solenodon. Venom and saliva was separated by HPLC and either directly analysis by HR FT MS/MS (Top-down) or further decomplexed by SDS-PAGE followed by in-gel trypsine digestion and HR LC-MS/MS analysis (bottom-up, Venom only). For shotgun bottom-up comparison of venom and saliva proteins, samples were directly reduced, alkylated, digested with trypsin and measured by HPLC-MS/MS. Additional bottom-up analysis was performed from bioactivity guided fractionation experiments.
Project description:We have used computational and experimental biology approaches to identify candidate mechanisms of action of a traditional Chinese medicine; Compound Kushen Injection (CKI), in a breast cancer cell line in which CKI has been shown to cause apoptosis. Because CKI is a complex mixture of plant secondary metabolites, we used a high-performance liquid chromatography (HPLC) fractionation and reconstitution approach to define chemical fractions required for CKI to induce apoptosis in MDA-MB-231 cells. Our initial fractionation separated major from minor compounds, and showed that the major compounds accounted for little of the activity of CKI. By systematically perturbing the major compounds in CKI we found that removal of no single major compound could alter the effect of CKI on cell viability and apoptosis. However, simultaneous removal of two major compounds identified oxymatrine and oxysophocarpine as critical compounds with respect to CKI activity. We then used RNA sequencing and transcriptome analysis to correlate compound removal with gene expression and phenotype data. We determined that many compounds in CKI are required for its effectiveness in triggering apoptosis but that significant modulation of its activity is conferred by a small number of compounds. In conclusion, CKI may be typical of many plant based extracts that contain many compounds in that no single compound is responsible for all of the bioactivity of the mixture and that many compounds interact in a complex fashion to influence a network containing many targets.
2019-05-29 | GSE125743 | GEO
Project description:Effects of crude extracts from Trichoderma simmonsii on Fusarium pseudograminearum
Project description:Screening a library of 573 cyanobacteria extracts for inhibition of the quorum sensing regulated prodigiosin production of Serratia marcescens, an extract of the cyanobacterium Fischerella ambigua (Näg.) Gomont 108b was found to drastically increase the prodigiosin production. Bioactivity-guided isolation of the active compounds resulted in the two new natural products ambigol D and E along with the known ambigols A and C. Ambigol C treatment increased prodiginine production of Serratia sp. ATCC 39006 (S39006) by a factor of 10, while ambigols A and D were found to have antibiotic activity against this strain. RNA-Seq of S39006 treated with ambigol C and subsequent differential gene expression and functional enrichment analyses indicated a significant downregulation of genes associated with the translation machinery and fatty acid biosynthesis in Serratia, as well as increased expression of genes related to the uptake of l-proline. These results suggest that the ambigols increase the prodiginine production in S39006 not by activating the SmaIR quorum sensing system, but possibly by increasing the precursor supply of l-proline and malonyl-CoA.
Project description:Traditional medicine and the use of herbal remedies are well established in the African health care system. For instance, Violaceae plants are used for antimicrobial or anti-inflammatory applications in folk medicine. This study describes the phytochemical analysis and bioactivity screening of four species of the violet tribe Allexis found in Cameroon. Allexis cauliflora, Allexis obonensis, Allexis batangae and Allexis zygomorpha were evaluated for the expression of circular peptides (cyclotides) by mass spectrometry. The unique cyclic cystine-rich motif was identified in several peptides of all four species. Knowing that members of this peptide family are protease inhibitors, the plant extracts were evaluated for the inhibition of human prolyl oligopeptidase (POP). Since all four species inhibited POP activity, a bioactivity-guided fraction approach was performed to isolate peptide inhibitors. These novel cyclotides, alca 1 and alca 2 exhibited IC50 values of 8.5 and 4.4 µM, respectively. To obtain their amino acid sequence information, combinatorial enzymatic proteolysis was performed. The proteolytic fragments were evaluated in MS/MS fragmentation experiments and the full-length amino acid sequences were obtained by de novo annotation of fragment ions. In summary, this study identified inhibitors of the human protease POP, which is a drug target for inflammatory or neurodegenerative disorders.