Identification of a Small Molecule That Inhibits the Interaction of LPS Transporters LptA and LptC

Abstract

The need for novel antibiotics has become imperative with the increasing prevalence of antibiotic resistance in Gram-negative bacteria in clinics. Acting as a permeability barrier, lipopolysaccharide (LPS) protects Gram-negative bacteria against drugs. LPS is synthesized in cells and transported to the outer membrane (OM) via seven lipopolysaccharide transport (Lpt) proteins (LptA–LptG). Of these seven Lpt proteins, LptC interacts with LptA to transfer LPS from the inner membrane (IM) to the OM, and assembly is aided by LptD/LptE. This interaction among the Lpt proteins is important for the biosynthesis of LPS; therefore, the Lpt proteins, which are significant in the assembly process of LPS, can be a potential target for new antibiotics. In this study, a yeast two-hybrid (Y2H) system was used to screen compounds that could block LPS transport by inhibiting LptA/LptC interaction, which finally disrupts the biosynthesis of the OM. We selected the compound IMB-0042 for this study. Our results suggest that IMB-0042 disrupts LptA/LptC interaction by binding to both LptA and LptC. Escherichia coli cells, when treated with IMB-0042, showed filament morphology, impaired OM integrity, and an accumulation of LPS in the periplasm. IMB-0042 inhibited the growth of Gram-negative bacteria and showed synergistic sensitization to other antibiotics, with low cytotoxicity. Thus, we successfully identified a potential antibacterial agent by using a Y2H system, which blocks the transport of LPS by targeting LptA/LptC interaction in Escherichia coli.

The Prediction of LptA and LptC Protein–Protein Interactions and Virtual Screening for Potential Inhibitors

“…Thereafter, 2000 ns MD simulations were conducted on IMB-881 and IMB-0042 with LptA or LptC complex. It can be observed that the two molecules underwent multiple cycles of dissociation and reassociation throughout the entire 2000 ns simulation (Figure 9) as well, which is consistent with their slow-binding-slow-dissociation kinetic characteristics obtained by surface plasmon resonance (SPR) [11,12,21].…”

“…Currently, two molecular inhibitors (IMB-881 and IMB-0042) (Figure 2) that target the LptA and LptC interaction have been reported, but the precise molecular mechanisms are currently unclear [12,13]. Another peptide inhibitor, Thanatin, targeting the interaction between LptA and LptC, was also reported [14,15], and the co-crystal structure of Thanatin and LptA was solved to indicate the binding pose (PDB ID: 6GD5) [16].…”

Thanksgiving to Yeast, the HMGB Proteins History from Yeast to Cancer

“…The Y3H is an extension of the Y2H but introducing a third hybrid component, usually a small molecule that can make possible or interfere with the protein-protein interaction, or a RNA molecule allowing the detection of protein-RNA interactions. All these methods allow the study of interactions between proteins of any other biological origin in the yeast model; for this reason, the Y2H has been extensively used in pharmacological screenings for novel drugs [23,24].…”