title: Structural and sequence analysis of class A β-lactamases with respect to avibactam inhibition: impact of Ω-loop variations.
abstract: There exists a significant diversity among class A β-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-β-lactam β-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.
  
  
  
  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.
  
  
  
  The majority of the key residues involved in avibactam binding were well conserved across the different sub-families of class A β-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 β-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.
  
  
  
  Overall, the computational and experimental analyses suggest that the vast majority of class A β-lactamases should be well inhibited by avibactam, although a very small number of outliers exist.
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, MA, USA slahiri@macrolide.com.
authors.forename: Sushmita D
authors.initials: SD
authors.lastname: Lahiri
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, MA, USA.
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Bradford
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, MA, USA.
authors.forename: Wright W
authors.initials: WW
authors.lastname: Nichols
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, MA, USA.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Alm
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D053961:Azabicyclo Compounds / Q000378:metabolism / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002442:Ceftazidime / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D007711:Klebsiella pneumoniae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017421:Sequence Analysis; D065093:beta-Lactamase Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D001618:beta-Lactamases / Q000737:chemistry* / Q000145:classification / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D053961:Azabicyclo Compounds; D001426:Bacterial Proteins; D065093:beta-Lactamase Inhibitors; C543519:avibactam; D002442:Ceftazidime; C108863:beta-lactamase PER-1; D001618:beta-Lactamases