Interactions of OXA-48 class β-lactamases with substrates and inhibitors

Abstract

The major resistance mechanism to the administration of β-lactam antibiotics in gram negative bacteria is the expression of β-lactamase enzymes. OXA-48 is a clinically concerning class D β-lactamase enzyme that is found wide-spread in Enterobacterales and worryingly confers resistance to a range of β-lactams, including the potent carbapenem class of antibiotics whose usage are typically reserved for only the most serious drug-resistant infections. Despite the extensive structural and biochemical information available on OXA-48, it still remains unclear how OXA-48-like enzymes inactivate carbapenem substrates, and also how their structural features define the potency of various β-lactamase inhibitors (BLIs). Therefore, using a combined experimental and computational structural biology approach, this thesis aims to describe novel interactions between OXA-48-like β-lactamases and carbapenem substrates as well as clinically established and investigational BLIs. Firstly, OXA-48 is structurally characterised in complex with the carbapenem antibiotics meropenem and ertapenem, capturing covalently bound acyl-enzyme intermediates that exhibit differing tautomeric states, with molecular dynamics simulations of these complexes revealing a preference for the Δ2-enamine tautomer to adopt a hydrolytically-active binding pose. The next chapter describes the structural characterisation of OXA-519, an OXA-48 natural variant with a single amino acid substitution that appears to enhance its activity against 1β-methyl carbapenems, yielding a novel binding mode for hydrolysed carbapenem-derivatives. Finally, this thesis summarises interactions between OXA-48 (and variants) and diazabicyclooctane (DBO) BLIs avibactam and nacubactam, as well as with mono-, biand tricyclic boronate compounds, and suggests how these interactions may relate to their varying inhibitory potencies.

Date of Award	1 Oct 2024
Original language	English
Awarding Institution	University of Bristol
Supervisor	Jim Spencer (Supervisor)