Antibiotic compositions and related screening methods

摘要

Moenomycin inhibits bacterial growth by clocking the transglycosylase activity of class A penicillin-binding proteins (PBPs), which are key enzymes in bacterial cell wall synthesis. The binding affinities of moenomycin A with various truncated PBPs were compared showing that the transmembrane domain is important for moenomycin binding. Full-length class-A PBPs from 16 bacterial species were produced, and their binding activities showed a correlation with the antimicrobial activity of moenomycin against Enterococcus faecalis and Staphylococcus aureus. Moreover, a fluorescence anisotropy-based high-throughput assay was developed and used successfully for identification of transglycosylase inhibitors.

主权项

1. A method for high throughput screening (HTS) of antibacterial agents, the method comprising:
providing a candidate agent for screening; providing a class A penicillin-binding protein, or fragment thereof (PBP), from a bacterial species, comprising at least a transmembrane (TM) domain and a transglycosylase (TG) domain; determining a first binding affinity of the candidate agent to the at least transmembrane (TM) domain and transglycosylase (TG) domain-containing fragment of the class A penicillin-binding protein; comparing the first binding affinity to a second binding affinity of the candidate agent to a class A penicillin-binding protein fragment that lacks the TM domain; identifying a candidate agent as a putative transglycosylase inhibitor of the bacterial species as the antibacterial agent, wherein the candidate agent exhibits a higher first binding affinity to the transmembrane (TM) domain-containing class A penicillin-binding protein, or fragment thereof, as compared to the second binding affinity to the PBP fragment that lacks the TM domain; and confirming the transglycosylase inhibitor activity of the putative transglycolase inhibitor by testing for an ability to inhibit growth of the same bacterial species whose PBP transmembrane domain-containing fragment it binds with higher affinity, wherein the binding affinity between the candidate agent and the transmembrane (TM) domain of the class A penicillin-binding protein is determined by anisotropy measurement assay.