Browsing by Author "Giard, Josephine Marie"

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    Data-driven discovery of novel lasso peptides with pharmaceutical applications
    (Heriot-Watt University, 2024-10) Giard, Josephine Marie; Kerr, Doctor Leena; Smith, Professor David; Alexander, Doctor Ross
    Antimicrobial resistance is a major global health threat and to combat it, there is a need to discover novel antimicrobials. A promising target for novel drug discovery are lasso peptides, a class of secondary metabolites with desirable characteristics and sought-after bioactivities. Here, novel lasso peptides with antimicrobial activity and Streptomyces origin were sought by using DNA sequencing data to guide experimental procedures. Initially, it was tested if a high abundance of Actinobacteria and Streptomyces can be used as an indicator for environments with high potential for secondary metabolism. Subsequently, metagenomic and bacterial whole genome sequencing data were assessed for the presence of lasso peptide gene clusters which led to the prediction of 177 potential lasso peptide gene clusters. Predicted lasso peptide gene clusters were further analysed for antimicrobial potential, novelty, sequence diversity, and Actinobacterial origin, and two lasso peptide gene clusters were chosen as cloning targets for further experiments. One target lasso peptide gene cluster was amplified from eDNA extracted from a volcanic cave biofilm and subsequently expressed by cell-free biosynthesis and maltose-binding protein cloning. The antimicrobial potential of the expressed novel lasso peptide was tested by screening for inhibitory activity against a selection of (multidrug resistant) ESKAPE pathogens. The results indicated an effect on the growth of Staphylococcus aureus. In addition to the expression of individual lasso peptide gene clusters, Streptomyces strains were isolated from environments suggested to be rich in lasso peptides. A selection of isolates was PCR-screened for lasso peptide genes. Positive PCR results could be confirmed by Sanger but not by whole genome sequencing.