Single-Molecule Force Spectroscopy and Imaging of the Vancomycin/d-Ala-d-Ala Interaction

Abstract

The clinically important vancomycin antibiotic inhibits the growth of pathogens such as Staphylococcus aureus by blocking cell wall synthesis through specific recognition of nascent peptidoglycan terminating in D-Ala-D-Ala. Here, we demonstrate the ability of single-molecule atomic force microscopy with antibiotic-modified tips to measure the specific binding forces of vancomycin and to map individual ligands on living bacteria. The single-molecule approach presented here provides new opportunities for understanding the binding mechanisms of antibiotics and for exploring the architecture of bacterial cell walls.

Analysis of membrane interactions of antibiotic peptides using ITC and biosensor measurements

“…These observation might be explained by the different membrane interactions of both peptides. Vancomycin targets the D-Ala-D-Ala motif of lipid II [13,14] and has not been described to interact with bacterial membranes, which is in good agreement with the hydrophilic polar structure of the glycopeptide (Figure 1). In contrast, gallidermin comprises hydrophobic moieties which allow for membrane interaction.…”

Interaction of an anticancer peptide fragment of azurin with p53 and its isolated domains studied by atomic force spectroscopy

“…To verify the presence of a monolayer and define its height, a bare tip was thus used to scratch the substrate, working in contact mode as described in Funari et al 27 (results not shown). With a functionalized tip thousands of force curves were thus recorded on such a substrate with an intermediate degree of coverage (condition also suggested by Gilbert et al), 35 at several distinct points in which a corresponding percentage of specific events were observed. The approach speed was kept constant while the retraction one, v, was varied from 50 to 4190 nm/s.…”

Recent progress in AFM molecular recognition studies

“…2) [11,[31][32][33]. To understand this behavior, one must recall that when viewing ligand-receptor binding on the singlemolecule level, the average lifetime of a ligand-receptor bond, τ(0), can be taken as the inverse of the kinetic off-rate constant, τ(0)=1/k off .…”

“…4) [33]. Fluorescence microscopy with a fluorescent vancomycin probe was used to visualize D-Ala-D-Ala sites of nascent peptidoglycan in the cell wall of dividing Lactococcus lactis cells (Fig.…”

“…Adhesion events are essentially located in the septum region (red line), more specifically on the ring-like structure, suggesting that newly formed peptidoglycan is inserted. Reprinted with permission from [33] composition on top of a topographical image and can detect compositional changes occurring during biological processes.…”