Identification of a Compound That Inhibits the Growth of Gram-Negative Bacteria by Blocking BamA-BamD Interaction.

Yan Li,Xiaohong Zhu,Yuan Lin,Minghua Chen,Ningyu Zhu,Xuefu You,Yanchang Wang,Shuyi Si,Jing Zhang

doi:10.3389/fmicb.2020.01252

Abstract

The demand for novel antibiotics is imperative for drug-resistant Gram-negative bacteria which causes diverse intractable infection disease in clinic. Here, a comprehensive screening was implemented to identify potential agents that disrupt the assembly of β-barrel outer-membrane proteins (OMPs) in the outer membrane (OM) of Gram-negative bacteria. The assembly of OMPs requires ubiquitous β-barrel assembly machinery (BAM). Among the five protein subunits in BAM, the interaction between BamA and BamD is essential for the function of this complex. We first established a yeast two-hybrid (Y2H) system to confirm the interaction between BamA and BamD, and then screened agents that specifically disrupt this interaction. From this screen, we identified a compound IMB-H4 that specially blocks BamA–BamD interaction and selectively inhibits the growth of Escherichia coli and other Gram-negative bacteria. Moreover, our results suggest that IMB-H4 disrupts BamA–BamD interaction by binding to BamA. Strikingly, E. coli cells having been treated with IMB-H4 showed impaired OM integrity and decreased the abundance of OMPs. Therefore, an antibacterial agent was identified successfully using Y2H system, and this compound likely blocks the assembly of OMPs by targeting BamA–BamD interaction in Gram-negative bacteria.

Highlights

Global health has been subjected in jeopardy as a result of emerging strains of drug-resistant Gram-negative bacteria (Wellington et al, 2013)
In the barrel assembly machinery (BAM) complex, the interaction between BamA and BamD is crucial for rapid integration of outer-membrane proteins (OMPs) to outer membrane (OM)
In AH109 cells, the transcription of three reporter genes ADE2, HIS3, and LacZ can be activated by the interaction between BamA and BamD, which can be validated by detecting β-gal activity after growing yeast cells on synthetic dropout (SD)/-Leu-Trp-Ade-His plate

Summary

Introduction

Global health has been subjected in jeopardy as a result of emerging strains of drug-resistant Gram-negative bacteria (Wellington et al, 2013). Very limited drugs are available for Gram-negative bacteria infection treatment (Lepore et al, 2019). The major hurdle for efficient elimination of infection is the presence of outer membrane (OM) in Gram-negative bacteria. The OM is a double-layer hydrophobic structure that envelopes the bacteria and functions as a highly selective permeability barrier, which facilitate bacteria with resistance to unfavorable surrounding environment and antibiotics, thereby deactivating many antibiotics prescriptions in the clinic (Nikaido, 2003). Antibiotic targeting the OM structure would have potential to kill Gram-negative bacteria or sensitize them to antibiotics unable to penetrate through the OM.

Methods

Results

Conclusion