Rhombohedral Trap for Studying Molecular Oligomerization in Membranes: Application to Daptomycin
Ming-Tao Lee1,2*, Wei-Chin Hung3, Huey W. Huang4
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
4Department of Physics and Astronomy, Rice University, Houston, TX, USA
* Presenter:Ming-Tao Lee, email:mtlee@nsrrc.org.tw
A persistent problem in the studies of membrane-active peptides, including antimicrobial peptides and pathogenic amyloidal peptides, is the lack of methods for investigating their molecular configurations in the membranes. These peptides spontaneously bind to membranes from solutions, and often form oligomers that induce changes of membrane permeability. For antimicrobials such actions appear to relate to the antimicrobial mechanisms, but for amyloidal peptides the oligomerization has been linked to neurodegenerative diseases. In many cases, no further understanding of such oligomerization problems has been made due to the lack of structural information. In this article we will demonstrate a method of trapping such peptide oligomers in a rhombohedral (R) phase of lipid so that the oligomers can be subjected to 3D diffraction analysis. The condition for forming the R phase and the electron density distribution in the rhombohedral unit cell provide information of peptide-lipid interaction and the molecular size of the trapped oligomer. Such information can not be obtained from membranes in the planar configuration. For illustration, we apply this method to daptomycin, an FDA-approved antibiotic that attacks membranes containing phosphatidylglycerol, in the presence of calcium ions. We have successfully used the brominated phosphatidylglycerol to perform bromine-atom anomalous diffraction in the rhombohedral phase containing daptomycin and calcium ions. The preliminary results apparently exhibit diffraction data related to daptomycin oligomers. We believe that this method will also be applicable to the difficult problems of amyloidal peptides, such as the amyloid beta of Alzheimer's disease.

Keywords: membrane-active peptides, oligomerization, rhombohedral phase, daptomycin, anomalous X-ray diffraction