Daptomycin Induces Transient Membrane-Permeabilization
Ming-Tao Lee1,2*, Wei-Chin Hung3, Meng-Hsuan Hsieh4, Hsiung Chen1, Yu-Yung Chang1, Huey W. Huang5
1Scientific Research Division, National Synchrotron Radiation Research Center, Hsinchu, Taiwan
2Department of Physics, National Central University, Jhongli, Taiwan
3Department of Physics, R. O. C. Military Academy, Fengshan, Kaohsiung, Taiwan
4Institute of Biotechnology, National Taiwan University, Taipei, Taiwan
5Department of Physics & Astronomy, Rice University, Houston, Texas, USA
* presenting author:Ming-Tao Lee, email:mtlee@nsrrc.org.tw
Daptomycin, a cyclic lipopeptide, represents a new structural class of the FDA approved antibiotics. The drug interacts with the cytoplasmic membranes of Gram-positive pathogens causing membrane permeabilization to ions and cell death. The antibiotic activity is calcium ion-dependent and correlates with the target membrane’s content of phosphatidylglycerol (PG). Fluorescence resonance energy transfer has detected daptomycin aggregation upon membrane binding and this has led to the hypothesis that daptomycin forms ion channels in the membrane. However many aspects of dayptomycin-membrane interactions are still unknown. Here we used X-ray scattering and CD analysis to clarify the physicochemical state of daptomycin in solution. With X-ray we determine that daptomycin in solution is monomeric at concentrations relevant to its therapeutic use. A careful CD analysis of daptomycin with Ca2+ and PG-containing membranes shows that there are only two states identifiable by CD, one before and one after membrane binding and the stoichiometry ratio of calcium to daptomycin is 3 to 2 in their binding reaction. To test the proposed daptomycin channels, we performed K+ and Ca2+ leakage experiment, comparing daptomycin with gramicidin, ionomycin, valinomycin and penetratin. The results revealed the property of transient ion conduction induced by daptomycin, dissimilar to gramicidin channel or ion carriers but similar to penetratin. Previous experiments have shown that penetratin does not form ion channels, but both daptomycin and penetratin exhibit a lipid extracting effect. The effect might create transient water pore defects that can explain the transient ion leakage induced by dayptomycin and penetratin.

Keywords: daptomycin, lipopeptide, X-ray scattering, CD, lipid extracting effect