Interaction Between Host Defence Peptides And Mycobacterial Membranes

Mycobacterium tuberculosis is resistant against the antimicrobial activity of a large number of host defence peptides (HDPs) in the phagosome, which are active against other Gram-negative or -positive bacteria. One reason for this resistance is the unique structure of the mycobacterial cell wall. Mycobacteria produce a thick mycolate-rich outer covering which functions as an exceptionally efficient barrier. We demonstrated that the membrane forming properties of trehalose dimycolate (TDM), as one of the important components of the mycobacterial outer barrier, are unique. TDM, which is organized in domains, leads to an increase of the overall membrane stability by a factor of three (as obtained in force spectroscopic experiments using atomic force microscopy). The membrane structure was investigated by X-ray reflectivity and GISAXS. Moreover, we used reconstituted planar lipid bilayers composed of TDM and phospholipids to perform electro-physiological experiments for determining the membrane permeabilizing activity of different HDPs. Furthermore, we used atomic force microscopy to characterize the surface topography of reconstituted membranes and mycobacteria. The biophysical results were compared with results from killing assays. Interestingly LL32, which is the highly active fragment of human cathelicidin, induced dramatic changes in the membrane morphology without inducing a significant permeabilization. LL32 is able to kill other Gram positive or negative bacteria by induction of membrane lesions, but up to a concentration of 30 μg/ml LL32 could not kill M. tuberculosis. Surprisingly, LL32 even stimulated an increased growth of the mycobacteria. In contrast two synthetic peptides based on LL32 and a linear version of the human beta-defensin-3 (hBD-3) showed a pronounced killing and permeabilization.