Browsing by Author "Clark, Trevor N."

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    Discovery of an isothiazolinone-containing antitubercular natural product Levesquamide
    (American Chemical Society, 2020-05-04) Liang, Libang; Haltli, Bradley; Marchbank, Douglas H.; Fischer, Maike; Kirby, Christopher W.; Correa, Hebelin; Clark, Trevor N.; Gray, Christopher A.; Kerr, Russell G.
    Antitubercular agent levesquamide is a new polyketide-nonribosomal peptide (PK-NRP) hybrid marine natural product isolated from Streptomyces sp. RKND-216. The structure contains a rare isothiazolinone moiety which has only been reported in collismycin SN. Structure elucidation by NMR spectroscopy was a significant challenge due to a deficiency of protons in this aromatic moiety. Therefore, the genome of Streptomyces sp. RKND-216 was sequenced to identify the levesquamide biosynthetic gene cluster (BGC). Analysis of the BGC provided structural insights and guided stable-isotope labeling experiments, which led to the assignment of the fused pyridine-isothiazolinone moiety. The BGC and the labeling experiments provide further insights into the biosynthetic origin of isothiazolinones. Levesquamide exhibited antimicrobial activity in the microplate alamarBlue assay (MABA) and low oxygen recovery assay (LORA) against Mycobacterium tuberculosis H37Rv with minimum inhibitory concentration (MIC) values of 9.65 and 22.28 µM, respectively. Similar activity was exhibited against rifampicin- and isoniazid-resistant M. tuberculosis strains with MIC values of 9.46 and 9.90 µM, respectively. This result suggests levesquamide has a different mode of action against M. tuberculosiscompared to the two first-line antitubercular drugs rifampicin and isoniazid. Furthermore, levesquamide shows no cytotoxicity against the Vero cell line, suggesting it may have a useful therapeutic window.
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    The application of NMR and LC-HRMS based prioritization strategies for the discovery of natural products by endophytic fungi from medicinal plants
    (University of New Brunswick, 2019) Clark, Trevor N.; Gray, Christopher
    Natural products have been an abundant source of diverse and complex chemical structures and endophytic fungi have proven to be a valuable source of natural products. This thesis explores the diversity of natural products produced by fungal endophytes isolated from the leaves of medicinal plants used by the Canadian First Nations. Natural product isolation projects were selected based on three different screening strategies: The first employed antimicrobial activity as a basis for project prioritization; The second approach used nuclear magnetic resonance (NMR) metabolomics to identify extracts with atypical profiles due to the presence of structurally unique metabolites; the third approach used tandem liquid chromatography - high-resolution mass spectrometry (LC-HRMS) based metabolomic analyses to reveal extracts that contain potentially novel natural products. The bioactivity directed prioritization approach, either on its own or in combination with NMR based metabolomics, led exclusively to the isolation of known natural products, although some of the bioactivities measured for these compounds had not been previously reported. NMR based metabolomics prioritization, when used alone, also led to the isolation of known natural products or new isomers of known natural products. Finally, the third prioritization method using LC-HRMS based metabolomics led to the isolation of two new natural products. These data suggest that prioritization using LC-HRMS based metabolomics alone is the most effective method for the isolation of novel natural products. Future work should focus on extracts that are highlighted by both LC-HRMS based metabolomics as well as bioactivity screening in an effort to discover novel antimicrobial structures.