Binding of neomycin and calcium to phospholipids and other anionic compounds

In order to assess the potential importance of different cellular binding sites for the adverse effects of aminoglycosides (i.e. oto- and nephrotoxicity) the binding of neomycin and calcium to phospholipids and other anionic cell constituents was assayed in vitro. Phospholipids demonstrated binding affinities that strongly favored neomycin (Km, 10 to 100 microM) over calcium (Km, 300 to 1,120 microM). Both neomycin and calcium showed strongest binding to lipids with monoester phosphate groups: phosphatidic acid, phosphatidylinositol 4-phosphate, and phosphatidylinositol 4,5-bisphosphate. The lipids bound 0.2 to 0.4 molecules of neomycin and 0.5 to 1 molecule of calcium, respectively, per lipid. Anionic non-lipid compounds such as melanin, gangliosides or chondroitin sulfate were ineffective competitors of neomycin binding to lipids. The results emphasize the importance of phospholipids as cellular binding sites for aminoglycosides. Furthermore, if one considers extra- and intracellular calcium and neomycin concentrations, the relative affinity of lipids for these two compounds suggests an explanation for both the reversible and the essentially irreversible toxic effects of the aminoglycosides.