Mechanism of uptake of deglucoteicoplanin amide derivatives across outer membranes of Escherichia coli and Pseudomonas aeruginosa

Teicoplanin is a glycopeptide antibiotic which is ineffective against gram-negative bacteria because of its inability to penetrate the outer membrane. Removal of the sugar residues and attachment of polyamines to carbon 63 yielded two dibasic deglucoteicoplanin amides, MDL 62,766 (766) and MDL 62,934 (934), with moderate MICs for Escherichia coli (2 to 4 micrograms/ml) and Pseudomonas aeruginosa (8 to 32 micrograms/ml) compared with those of the monobasic teicoplanin aglycone (16 and > 1,024 micrograms/ml, respectively). MICs were increased 16- to 32-fold by Mg2+ supplementation of Luria broth but not by Na+ supplementation at an equivalent ionic strength. Both 766 and 934 were capable of binding to P. aeruginosa lipopolysaccharide (LPS) at Mg(2+)-binding sites, as assessed by dansyl polymyxin displacement experiments. Furthermore, both compounds increased E. coli and P. aeruginosa outer membrane permeability to the hydrophobic fluorescent probe 1-N-phenylnaphthylamine (NPN), whereas the parent compounds teicoplanin aglycone and teicoplanin and the beta-lactam ceftazidime were totally ineffective. Addition of 1 mM Mg2+ blocked the increase in outer membrane permeability. Compound 766 had a lower MIC than 934 for both bacteria tested, bound to LPS with a higher affinity, and permeabilized outer membranes to NPN at lower concentrations. We propose that both deglucoteicoplanin amides exhibit increased gram-negative activity by virtue of their ability to access the self-promoted uptake pathway across the outer membrane.