Thesis

Decipher, disarm and disengage : understanding the biosynthesis and self-resistance mechanisms of kirromycin-like, elfamycin producing, streptomyces.

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Awarding institution
  • University of Strathclyde
Date of award
  • 2025
Thesis identifier
  • T17310
Person Identifier (Local)
  • 202085988
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Qualification Name
Department, School or Faculty
Abstract
  • Elfamycin antibiotics exhibit activity against Gram-positive bacteria by inhibiting translation via elongation factor EF-Tu. Elfamycins are characterised by mode of action rather than chemical structure, though minor structural modifications in these antibiotics can significantly alter their biological activity. Aurodox, a kirromycin-like elfamycin, was shown to inhibit the Type III Secretion System (T3SS) of Enteropathogenic (EPEC) and Enterohemorrhagic (EHEC) Escherichia coli along with effective anti-virulence treatment. This study aimed to investigate the diversity of kirromycin-like elfamycin biosynthetic gene clusters (BGCs), their evolution, resistance mechanisms, and how biosynthesis may be manipulated to create modified elfamycin variants which many be active against the T3SS of EHEC. Genome sequencing of the kirromycin producer Streptomyces ramocissimus was performed, and the BGC responsible for kirromycin production through comparison to the BGC found in Streptomyces collinus. Comparative analyses between kirromycin-like BGCs revealed the potential for natural variation in producing identical compounds via different genetic pathways. Additionally, a kirromycin-like BGC was identified in Streptomyces ISL094, which contained an additional methyltransferase on the BGC, similar to the aurodox BGC of S. goldiniensis, suggesting it could be an aurodox producer. Moreover, aurodox and kirromycin were found to downregulate the T3SS of EHEC, a trait previously thought to be unique to aurodox. Key genes within the aurodox BGC were manipulated and the BGC heterologously expressed to infer function of their roles in aurodox biosynthesis, where their derivatives showed similar effects on the T3SS. The protein, AurM*, thought to methylate the kirromycin molecule creating aurodox was assayed for methyltransferase activity in vitro, but activity was not demonstrated on kirromycin as a substrate. Finally, a potential "moonlighting" role for EF-Tu proteins is suggested, indicating that they might have additional functions related to T3SS regulation, where the knockdown phenotype of aurodox on the T3SS of EHEC was reversed when EF-Tu was expressed in EHEC.
Advisor / supervisor
  • Hoskisson, Paul A.
  • Burley, Glenn A.
Resource Type
DOI
Date Created
  • 2024

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