ナノメディシンとナノテクノロジーのジャーナル

概要

Molecular Addressability of Lipid Membrane Embedded Calixarenes towards Cytochrome C

Alexandra Poturnayova, Michael Leitner, Maja Snejdarkova, Peter Hinterdorfer, Tibor Hianik and Andreas Ebner

By means of the atomic force microscopy techniques we studied the surface topography of the supported bilayer lipid membranes (sBLM) composed of 1,2-sn-glycerodimyristoylfosfatidylcholine (DMPC) with incorporated calixarenes tOct [6]CH2COOH (CX) specific to cytochrome c (cyt c). It is supposed that cyt c interacts with CX through amino groups of lysine residues at its surface. Therefore we also applied single molecule force spectroscopy (SMFS) to analyze the mechanisms of interaction of cyt c with the CX. In later case cyt c or individual NH2 group have been connected to the AFM tip through special linker. The topography of bare sBLM in a gel state (T=19ºC) revealed relatively smooth surface (RRMS=0.18 nm) and thickness ~5.1 nm which agrees well with previous studies. Incorporation of CX into DMPC bilayer resulted in increase of the surface roughness (RRMS= 0.39 nm) and in increase of thickness in average by 0.5 nm. The incubation of the layer with30 nM of cyt c resulted in a surface smoothing (RRMS=0.32 nm) and in a further increase of the thickness between 0.7 to 1.2 nm. The SMFS experiments with cyt c modified AFM tips approved its specific binding to CX and allowed us to determine the binding parameters koff (1.14 ± 0.59 s-1) and xβ (3.98 ± 0.63Å). SMFS experiments with an amino-ended linker also resulted in highly specific interactions with comparable values for koff (2.74 ± 0.66 s-1) and xβ (5.91 ± 2.55 Å). This suggests that both electrostatic and amino group specific interactions between cyt c and CX cavity exist.