Scotti, F; (2017) Novel potential peptide therapeutics for tuberculosis therapy. Doctoral thesis , UCL (University College London).

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Abstract

Despite the existence of vaccinations, diagnostic tools and treatments, tuberculosis (TB) prevalence is increasing because of the circulation of people and misuse of antibiotics, giving rise to growing numbers of drug resistant strains of Mycobacterium tuberculosis. There is therefore a pressing need to look for new strategies against TB, in the hope of finding new drugs with novel mechanisms of anti-tubercular action or ways to potentiate the activity of already existing drugs and reduce treatment duration. This thesis explores the employment of peptides in anti-tuberculosis therapy. The project was initiated by the identification of a novel therapeutic target in M. tuberculosis: murein peptide ligase (Mpl, Rv3712), an enzyme involved in the bacterial peptidoglycan recycling process. The aim is to synthesise its putative natural substrates (peptidoglycan peptide fragments) to characterise its activity and synthesise sequence analogues. These analogues were tested on the whole-cell and will be evaluated for inhibitory activity on the recombinant Mpl enzyme and eventually could be used in combination with existing or new drugs to see whether they increase anti-tubercular potency and thus combat resistance. Attainment of the putative substrate required the synthesis of mDAP, an unusual amino acid unique to peptidoglycan. Its synthesis was successfully completed and it was incorporated in the tripeptide Mpl putative substrate. Solid-phase synthesis has been used successfully and proved effective for rapid synthesis of multiple short peptide analogues in parallel. In addition it was used to synthesise anti-tuberculosis lasso peptides, lariatins A and B, lassomycin and analogues, to evaluate the structural requirements for biological activity. The method for the heterologous expression and purification of recombinant Mpl from M. tuberculosis has been confirmed as successful, and the enzyme is available for future target-based evaluation using the synthesised mDAP-containing tripeptide and eventually for other mDAP-containing PG fragments and analogues.

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