Browsing by Author "Rowland Dugald Joseph"

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    Heterologous expression of a putative lodAB operon derived from thalassomonas viridans
    (University of the Western Cape, 2024) Rowland Dugald Joseph
    The growing incidence of antibiotic resistance in bacteria to current antimicrobial compounds is a major concern worldwide. Identifying novel antimicrobial compounds is vital to mitigate the increasing prevalence of antibiotic-resistant bacteria and other pathogens. One of the methods used to find novel antimicrobial compounds is genome mining. The genome mining of marine organisms, which have been poorly represented in research, may reveal a potential source of new antimicrobial compounds which can be used as antibiotics. This study focused on the characterisation of an antibacterial protein identified by the genome mining of a marine microorganism. Thalassomonas viridans is a microorganism that is known to produce antibiotic secondary metabolites (Adams, 2019). The genomic annotation of the T. viridans genome indicates that there are two sequences with similarity to the lodA gene that codes for l-lysine ε-oxidase (LodA) - first described as an antimicrobial protein produced by the marine bacteria Marinomonas mediterranea. LodA is an extracellular enzyme that has antibacterial properties associated with the production of hydrogen peroxide (H2O2) as a product of lysine oxidation (Gómez D et al.;2006). The lodA sequences submitted to a wider BLAST analysis showed that similar sequences are present in the genomes of a number of other microorganisms. In this study the heterologous expression of the lodA genes from T. viridans was conducted in Escherichia coli to produce, purify, and characterise the proteins. Multiple methods of E.coli expression to produce the protein were explored, and multiple activity assays wereemployed to characterise the proteins produced. The LodA protein was produced at low concentrations and no activity was detected on a wide range of amino acid substrates