Structural Insights into Inhibition of Escherichia coli Penicillin-binding Protein 1B

Abstract

Edited by F. Peter GuengerichIn Escherichia coli, the peptidoglycan cell wall is synthesized by bifunctional penicillin-binding proteins such as PBP1b that have both transpeptidase and transglycosylase activities. The PBP1b transpeptidase domain is a major target of ␤-lactams, and therefore it is important to attain a detailed understanding of its inhibition. The peptidoglycan glycosyltransferase domain of PBP1b is also considered an excellent antibiotic target yet is not exploited by any clinically approved antibacterials. Herein, we adapt a pyrophosphate sensor assay to monitor PBP1b-catalyzed glycosyltransfer and present an improved crystallographic model for inhibition of the PBP1b glycosyltransferase domain by the potent substrate analog moenomycin. We elucidate the structure of a previously disordered region in the glycosyltransferase active site and discuss its implications with regards to peptidoglycan polymerization. Furthermore, we solve the crystal structures of E. coli PBP1b bound to multiple different ␤-lactams in the transpeptidase active site and complement these data with gel-based competition assays to provide a detailed structural understanding of its inhibition. Taken together, these biochemical and structural data allow us to propose new insights into inhibition of both enzymatic domains in PBP1b.

Potential of 2-Chloro-N-(4-fluoro-3-nitrophenyl)acetamide Against Klebsiella pneumoniae and In Vitro Toxicity Analysis

“…With PBP1, the A2 and A1 molecules performed hydrogen bonding interactions with the Thr701, Thr 699, Asn574, and Ser510 residues, together with several Van der Walls interactions with other residues from the active site, as can be seen in Figure 2 . However, the presence of chloro in A2 is fundamental to better stabilize this molecule in the active site of the enzyme, due to Van der Walls interactions with the Ser510 and Thr702 residues that are important for a good anchorage in this active site [ 19 ]. These differences in anchoring reflect the potency of the antibacterial activity that was observed previously, confirming that the A2 molecule has a more pronounced antibacterial effect.…”

“…The chemical structures of the compounds were designed using the software MarvinSketch 18.5, their energies minimized in the program Hyperchem v. 8.0.3, using the molecular mechanics method (MM +) and the semi-empirical method AM1 (Austin Model 1) [ 36 ]. The enzymes were obtained from the Protein Data Bank ( ) together with their co-crystallized inhibitors, with PDB ID: 2VF52 (2.9 Å) [ 37 ], 5V3D3 (1.54 Å) [ 38 ], 5HLA4 (1.7 Å) [ 19 ], 2ZD85 (1.05 Å) [ 39 ], 3PBS6 (2.00 Å) [ 40 ], 6C3U7 (1.85 Å) [ 41 ], 1AJ68 (2.30 Å) [ 42 ], 1S149 (2.00 Å) [ 43 ]. Molecular docking was performed at Molegro Virtual Docker (MVD) software (v 6.0.1, Molegro ApS, Aarhus, Denmark), using the standard parameters of the software, the water molecules were removed and a template was generated in the co-crystallized inhibitor of the PDB.…”

Coupling of polymerase and carrier lipid phosphatase prevents product inhibition in peptidoglycan synthesis

“…In order to better understand the interaction of PgpB and PBP1B, we performed an in silico docking study using the published crystal structures of PgpB ( Tong et al, 2016 ) and PBP1B ( King et al, 2017 ) ( Fig. 8 ).…”

“…The docking models of PgpB-PBP1B complex were built using HADDOCK2.2 data-driven docking protocols ( Dominguez et al, 2003 ) and CNS1.2 ( Brünger et al, 1998 ) for the structure calculations. The initial coordinates of E. coli PgpB (PDB code 5JWY ( Tong et al, 2016 )) and E. coli PBP1B (PDB code 5HLD ( King et al, 2017 )) were used and the docking energy minimization occurred with a minimal conformational rearrangement of the partners. The multi domain docking was driven with ambiguous interaction restraints between the transmembrane helix of PBP1B (active residues 74–96) and PgpB (passive residues 1–235).…”

Modified Penicillin Molecule with Carbapenem-Like Stereochemistry Specifically Inhibits Class C β-Lactamases

“…8a). We also generated a model of MPC-1 in complex with the penicillin-binding (PB) domain of E. coli PBP1b (PDB code 5HL9) after superimposing the PB domain onto our P99 structure (29). Although the large number of PBPs identified in nature have functionally and structurally distinct N-terminal domains, their C-terminal PB domain is highly conserved (30).…”