Towards discovery of inhibitors of the undecaprenyl-pyrophosphate phosphatase BacA by virtual high-throughput screening

Marko Jukič; Rodolphe Auger; Victor Folcher; Matic Proj; Hélène Barreteau; Stanislav Gobec; Thierry Touzé

doi:10.1016/j.csbj.2022.05.010

Towards discovery of inhibitors of the undecaprenyl-pyrophosphate phosphatase BacA by virtual high-throughput screening

Comput Struct Biotechnol J. 2022 May 11:20:2360-2371. doi: 10.1016/j.csbj.2022.05.010. eCollection 2022.

Authors

Marko Jukič^{1

2}, Rodolphe Auger³, Victor Folcher³, Matic Proj⁴, Hélène Barreteau³, Stanislav Gobec⁴, Thierry Touzé³

Affiliations

¹ University of Maribor, Faculty of Chemistry and Chemical Engineering, Laboratory of Physical Chemistry and Chemical Thermodynamics, Smetanova 17, Maribor SI-2000, Slovenia.
² University of Primorska, Faculty of Mathematics, Natural Sciences and Information Technologies, Glagoljaška 8, Koper SI-6000, Slovenia.
³ Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Gif-sur-Yvette FR-91198, France.
⁴ Univerza v Ljubljani, Fakulteta za Farmacijo, Aškerčeva cesta 7, Ljubljana SI-1000, Slovenia.

Abstract

Increasing resistance to common antibiotics is becoming a major challenge that requires the development of new antibacterial agents. Peptidoglycan is an essential heteropolymer of the bacterial envelope that ensures the integrity and shape of all bacteria and is also an important target for antibiotics. The biosynthesis of peptidoglycan depends on a lipid carrier, undecaprenyl phosphate. As a byproduct of peptidoglycan polymerization, the lipid carrier is released as undecaprenyl pyrophosphate, which must be recycled to allow new polymerization cycles. To this end, it undergoes a dephosphorylation process catalyzed by the membrane phosphatase BacA, which is specific and highly conserved in bacteria. In the present study, we identified small molecules displaying inhibitory potency towards BacA. We began by preparing a commercial compound library, followed by high-throughput virtual screening by ensemble docking using the 3D structure of BacA and molecular dynamics snapshots to account for the flexibility of the protein. Of 83 compounds computationally selected and tested in a biochemical assay, one sulfamoylthiophene molecule showed significant inhibition of the undecaprenyl pyrophosphate dephosphorylation activity catalyzed by BacA. Subsequently, an additional 33 scaffold analogs were selected and acquired, of which 6 compounds exhibited BacA inhibition. The IC₅₀ values of these compounds ranged from 42 to 366 μM. In addition, significant antibacterial activity against Escherichia coli was observed in TolC/PAP2-depleted strains. We believe that the overall strategy followed in this study and the identified class of inhibitors provide a solid foundation for the further development of potent BacA-targeted inhibitors and the discovery of novel antibacterial compounds.

Keywords: Antibacterial drug design; Antibacterials; BacA; Binding site identification; C15-PP, Farnesyl pyrophosphate; C5-PP, Isopentenyl pyrophosphate; C55-P, Undecaprenyl phosphate; C55-PP, Undecaprenyl pyrophosphate; Ensemble docking; Escherichia coli; GlcNAc, N-acetylglucosamine; HTVS, High-Throughput Virtual Screening; In silico drug design; MD, Molecular Dynamics; MIC, Minimum Inhibitory Concentration; Molecular dynamics; MurNAc, N-acetylmuramic acid; Non-covalent inhibitors; PBPs, Penicillin-binding proteins; PG, Peptidoglycan; PP, Pyrophosphate; RA, Residual activity; RMSD, Root-mean-square deviation; TLC, Thin layer chromatography; Undecaprenyl pyrophosphate phosphatase; VS, Virtual Screening; Virtual screening.