Structural and sequence analysis of class A ß-lactamases with respect to avibactam inhibition: impact of Ω-loop variations.

BACKGROUND: There exists a significant diversity among class A ß-lactamases and the proliferation of these enzymes is a significant medical concern due to the ability of some members to efficiently hydrolyse both extended-spectrum cephalosporins and carbapenems. Avibactam is a novel non-ß-lactam ß-lactamase inhibitor that, in combination with ceftazidime, has recently obtained regulatory approval in the USA. Although avibactam is known to efficiently inhibit key class A enzymes, the diversity of this enzyme family warranted a more complete investigation to understand the breadth of the potential spectrum of inhibition. METHODS: Using the known residues critical for avibactam binding, a thorough structural and sequence-based conservation analysis was performed across >650 class A enzymes. Several variations that had the potential to impact avibactam inhibition were observed and representative enzymes were cloned and expressed isogenically to evaluate the impact of these variations. RESULTS: The majority of the key residues involved in avibactam binding were well conserved across the different sub-families of class A ß-lactamases, although some differences were observed. The differences in the Ω-loop of PER enzymes were found to impact the ability of avibactam to effectively protect ß-lactams against hydrolysis. However, substitutions in a key hydrogen-bonding residue (N170) in some of the GES variants were found to not have a significant impact on avibactam inhibition. CONCLUSIONS: Overall, the computational and experimental analyses suggest that the vast majority of class A ß-lactamases should be well inhibited by avibactam, although a very small number of outliers exist.