Class C β-Lactamases: Molecular Characteristics

Abstract

Class C β-lactamases or cephalosporinases can be classified into two functional groups (1, 1e) with considerable molecular variability (≤20% sequence identity). These enzymes are mostly encoded by chromosomal and inducible genes and are widespread among bacteria, including Proteobacteria in particular. Molecular identification is based principally on three catalytic motifs (⁶⁴SXSK, ¹⁵⁰YXN, ³¹⁵KTG), but more than 70 conserved amino-acid residues (≥90%) have been identified, many close to these catalytic motifs. Nevertheless, the identification of a tiny, phylogenetically distant cluster (including enzymes from the genera Legionella, Bradyrhizobium, and Parachlamydia) has raised questions about the possible existence of a C2 subclass of β-lactamases, previously identified as serine hydrolases. In a context of the clinical emergence of extended-spectrum AmpC β-lactamases (ESACs), the genetic modifications observed in vivo and in vitro (point mutations, insertions, or deletions) during the evolution of these enzymes have mostly involved the Ω- and H-10/R2-loops, which vary considerably between genera, and, in some cases, the conserved triplet ¹⁵⁰YXN. Furthermore, the conserved deletion of several amino-acid residues in opportunistic pathogenic species of Acinetobacter, such as A. baumannii, A. calcoaceticus, A. pittii and A. nosocomialis (deletion of residues 304-306), and in Hafnia alvei and H. paralvei (deletion of residues 289-290), provides support for the notion of natural ESACs. The emergence of higher levels of resistance to β-lactams, including carbapenems, and to inhibitors such as avibactam is a reality, as the enzymes responsible are subject to complex regulation encompassing several other genes (ampR, ampD, ampG, etc.). Combinations of resistance mechanisms may therefore be at work, including overproduction or change in permeability, with the loss of porins and/or activation of efflux systems.

Keywords: AmpC β-lactamases; ESAC; cephalosporinases; extended-spectrum; phylogeny; primary structure.

FIG 1

(A) Phylogram for representative and putative class C β-lactamases, compared with β-lactamases from classes A, B, and D. (B) Focused view on the phylogram of class C β-lactamases. The protein sequences of representative enzymes are listed in references , , , and . The sequences were filtered using CD-HIT (https://github.com/weizhongli/cdhit) at 90% sequence identity, then aligned with Clustal Omega (319). The tree was constructed using RAxML (320), and the phylogram generated using FigTree (version 1.4.3). The tree was unrooted.

FIG 2

Representative three-dimensional structure for class C β-lactamases. The structure of E. hormaechei P99 (formerly known as E. cloacae P99; PDB code 1BLS) (87) is colored in orange (α-helixes) and purple (β-strands). The Ω- and R2-loops are colored in green and blue, respectively. The most conserved residues (see Table 3) are represented as sticks and colored in cyan. The numbering of residues follows the SANC nomenclature (25).

FIG 3

Consensus partial protein sequences of species or their progenitors susceptible to expand their spectrum of inactivation. For E. coli, the consensus sequence was calculated from protein sequences of clusters A, B, C and D (32, 46, 50, 63). Residue boxed in yellow indicates 100% conserved. Underlined positions indicate at least two different residues (polymorphism).