Structures and ligand interactions of penicillin binding proteins in gram-negative bacteria

Abstract

Antibiotic resistance has become one of the major threats to global health, because of the increasing emergence of multi-drug resistant bacteria and the lack of new effective antibiotics to counteract them. Therefore, new antibiotics need to be developed, and bacterial cell wall biosynthesis has long been recognised as an excellent target for antibacterial discovery. Transpeptidation is the last reaction in the synthesis of peptidoglycan and is catalysed by penicillin binding proteins, the enzymes targeted by β-lactam antibiotics. In this project, a novel crystal structure of PBP2 has been determined from Acinetobacter baumannii, a Gram-negative pathogen commonly causing nosocomial infections, to aid the structure-based design of new PBP-targeting molecules. Structures of PBP2 in complex with b-lactam antibiotics and diazabicyclooctane compounds have been also generated, highlighting particular protein-ligand interactions that could inform the optimisation of PBP inhibitors from existing and novel chemical classes. Moreover, the structure of A. baumannii PBP2 has disclosed distinctive features in the transpeptidase domain of this enzyme, including a new zinc-binding site proximal to the catalytic pocket, that is essential for the cell wall elongation in this pathogen. β-lactam molecules generated by the polymerisation of ampicillin in solution have been also investigated, with particular emphasis on the interactions with PBPs and β-lactamases. The covalent binding of ampicillin and its polymers to various PBPs from Gram-negative bacteria have been confirmed by mass spectrometry, and structural studies of Pseudomonas aeruginosa PBP1a and PBP3 have been undertaken to elucidate the binding mode of these particular molecules. Ampicillin and ampicillin polymers are hydrolysed by the β-lactamase AmpC at different rates, and they also show differences in antimicrobial activity. This study provides the structural basis for the inhibition of important HMW-PBPs by β-lactam and non-β-lactam compounds, and it could be useful for the development of potent PBP inhibitors