Potentiation of Antibiotics against Gram-Negative Bacteria by Polymyxin B Analogue SPR741 from Unique Perturbation of the Outer Membrane

Therapeutics
targeting Gram-negative bacteria have the challenge of overcoming
a formidable outer membrane (OM) barrier. Here, we characterize the
action of SPR741, a novel polymyxin B (PMB) analogue shown to potentiate
several large-scaffold antibiotics in Gram-negative pathogens. Probing
the surface topology of Escherichia coli using atomic
force microscopy revealed substantial OM disorder at concentrations
of SPR741 that lead to antibiotic potentiation. Conversely, very little
cytoplasmic membrane depolarization was observed at these same concentrations,
indicating that SPR741 acts predominately on the OM. Truncating the
lipopolysaccharide (LPS) core with genetic perturbations uniquely
sensitized E. coli to SPR741, suggesting that LPS
core residues keep SPR741 at the OM, where it can potentiate a codrug,
rather than permit its entry to the cytoplasmic membrane. Further,
a promoter activity assay revealed that SPR741 challenge induced the
expression of RcsAB, a stress sensor for OM perturbation. Together,
these results indicate that SPR741 interacts predominately with the
OM, in contrast to the dual action of PMB and colistin at both the
outer and cytoplasmic membranes.