Project description:Untargeted-metabolomics LC-MS/MS analysis of commercial natural products pool, analyzed with different DDA settings with the objective to find the best one.
Project description:SDF-1 has been reported to trigger ADAMTS4,5 overexpression through activating CXCR4 signaling in chondrocytes. Here we described the transcriptional changes of SDF-1-treatment as well as natural products CXCR4 antagonists treatment.
Project description:We integrated quantitative proteomics and activity-based protein profiling (ABPP) approach to systematically elucidate the involved pathways and the covalent targets of natural products.
Project description:Anaerobic fungi (class Neocallimastigomycetes) thrive as low-abundance members of the herbivore digestive tract. The genomes of anaerobic gut fungi are poorly characterized and have not been extensively mined for the biosynthetic enzymes of natural products such as antibiotics. Here, we investigate the potential of anaerobic gut fungi to synthesize natural products that could regulate membership within the gut microbiome. Complementary 'omics' approaches were combined to catalog the natural products of anaerobic gut fungi from four different representative species: Anaeromyces robustus (A. robustus), Caecomyces churrovis (C. churrovis), Neocallimastix californiae (N. californiae), and Piromyces finnis (P. finnis). In total, 146 genes were identified that encode biosynthetic enzymes for diverse types of natural products, including nonribosomal peptide synthetases and polyketide synthases. In addition, N. californiae and C. churrovis genomes encoded seven putative bacteriocins, a class of antimicrobial peptides typically produced by bacteria. During standard laboratory growth on plant biomass or soluble substrates, 26% of total core biosynthetic genes in all four strains were transcribed. Across all four fungal strains, 30% of total biosynthetic gene products were detected via proteomics when grown on cellobiose. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) characterization of fungal supernatants detected 72 likely natural products from A. robustus alone. A compound produced by all four strains of anaerobic fungi was putatively identified as the polyketide-related styrylpyrone baumin. Molecular networking quantified similarities between tandem mass spectrometry (MS/MS) spectra among these fungi, enabling three groups of natural products to be identified that are unique to anaerobic fungi. Overall, these results support the finding that anaerobic gut fungi synthesize natural products, which could be harnessed as a source of antimicrobials, therapeutics, and other bioactive compounds. See publication: https://www.doi.org/10.1073/pnas.2019855118.
This research was performed under the Facilities Integrating Collaborations for User Science (FICUS) program (proposal:https://doi.org/10.46936/fics.proj.2018.50386/60000039) and used resources at the DOE Joint Genome Institute (https://ror.org/04xm1d337) and the Environmental Molecular Sciences Laboratory (https://ror.org/04rc0xn13), which are DOE Office of Science User Facilities operated under Contract Nos. DE-AC02-05CH11231 (JGI) and DE-AC05-76RL01830 (EMSL).
Project description:Finding the targets of natural products is of key importance in both chemical biology and drug discovery, and deconvolution of cofactor interactomes contribute to the functional annotation of the proteome. Identifying the proteins that underlie natural compound activity in phenotypic screens help to validate the respective targets and, potentially, expand the druggable proteome. Here, we present a generally applicable protocol for the photoactivated immobilisation of unmodified and microgram quantities of natural products on diazirine-decorated beads and their use for systematic affinity-based proteome profiling. We show that amongst 31 molecules of very diverse reported activity and biosynthetic origin, 25 could indeed be immobilised. Dose-response competition binding experiments using lysates of human or bacterial cells followed by quantitative mass spectrometry recapitulated <100 µM targets of 9 molecules. Among them, immobilisation of coenzyme A produced a tool to interrogate proteins containing a HotDog domain. Surprisingly, immobilisation of the cofactor flavin adenine dinucleotide (FAD) led to the identification of nanomolar interactions with dozens of RNA-binding proteins.