Lipoteichoic acid as a new target for activity of antibiotics: mode of action of daptomycin (LY146032)

Abstract

Daptomycin at the MIC allowed the cell mass increase of enterococcal strains and Bacilus subtilis to continue for 2 to 3 h at rates comparable to those of the controls. During this time the cell shape of the former changed to a rod configuration and that of the latter changed to long rods. In these bacteria, in which cell mass continued to increase, the MIC of daptomycin inhibited peptidoglycan synthesis by no more than 20% after 20 min of incubation and by roughly 50% after 2 h of incubation. Other macromolecules, such as DNA, RNA, and proteins, were only slightly affected. In contrast, incorporation of [14C]acetate into lipids was reduced by about 50% in the various strains after 20 min of treatment with daptomycin at the MIC. When the effect of the major lipid-containing polymers on synthesis was evaluated in detail, it was found that under conditions in which peptidoglycan and the other macromolecules mentioned above were inhibited only slightly (20%) and total lipid synthesis was inhibited by 50%, synthesis of teichoic and lipoteichoic acids was inhibited by 50 and 93%, respectively. Daptomycin was not found to enter the cytoplasm of either bacterial or mammalian cells. It bound, in the presence of calcium ions only, to whole bacterial cells, cell walls (both those that contained and those that did not contain membranes), and isolated membranes of bacterial and mammalian cells. Washing with EDTA removed daptomycin from all cells mentioned above and cell fractions except the bacterial membrane. It is concluded that lipoteichoic acid is most likely the primary target of daptomycin.Daptomycin (LY146032) is an acidic lipopeptide antibiotic that is active against gram-positive bacteria (F. T. Counter,

The action mechanism of daptomycin

“…58 Cell fractionation experiments detected daptomycin bound to cell walls and cell membranes, but not to cytosolic constituents. 59 The latter study also reported that daptomycin inhibits the biosynthesis of cell wall macromolecules, and the authors proposed inhibition of lipoteichoic acid biosynthesis as daptomycin's major mode of action. Several studies have attempted to determine whether daptomycin indeed directly blocks some enzyme or substrate in cell wall biosynthesis, or alternatively, inhibition of macromolecular synthesis occurs downstream of membrane permeabilization and loss of key metabolites; this question remains controversial (see section 11).…”

“…Elongated forms of Enterococcus and Bacillus cereus cells grown at MIC were reported in early studies. 59 Bacillus subtilis cells grown with daptomycin at below MIC appeared bent and elongated. 72 While elongated cell forms suggest that division is inhibited, the observation of additional, aberrant septa in Staphylococcus aureus 76 could mean that cell division may be dysregulated rather than disrupted.…”

“…However, their reported kinetics of cell killing and biosynthetic collapse differ only by some minutes. 79 Moreover, since daptomycin is tightly bound to the cell membrane, 59 its effect will not be promptly terminated by dilution, and thus the time course of its bactericidal action cannot be accurately determined. Therefore, no substantial evidence contradicts the tenet that indeed membrane permeabilization sets the pace for bactericidal action.…”

“…59 Such a preferential inhibition might arise from a requirement of LTA synthesis for some specific metabolite or ion that is lost particularly rapidly after permeabilization, and therefore does not prove a direct inhibition of the pathway.…”

Liposomal co-delivery of daptomycin and clarithromycin at an optimized ratio for treatment of methicillin-resistantStaphylococcus aureusinfection

“…The possible bactericidal mechanisms of daptomycin have been exhaustively investigated, especially the interactions between daptomycin and lipid membranes (Canepari et al, 1990;Silverman et al, 2001). It was reported that daptomycin-induced membrane permeabilization and depolarization, finally destroying the bacterial cell membrane, resulting in release of cellular contents (Canepari et al, 1990;Silverman et al, 2001).…”