Selective Organization of an α-Helical Hydrophobic Polypeptide/Pigment Complex in a Lipid Domain of Binary Lipid Bilayers

Abstract

We present here a novel approach to organization of an α-helical hydrophobic polypeptide complex in lipid bilayers in a 'lipid-domain-selective' manner. The key strategy is to use disulfide linkage between a cysteine-bearing polypeptide and a disulfide-based phospholipid, the latter of which forms a phase-separated lipid domain in a binary lipid bilayer with an incompatible phospholipid lacking disulfide group. The cross-linkage between the polypeptide and the phospholipid can be expected that the polypeptide acquires a higher affinity to the disulfide-based lipid domain than to the other one, resulting in selective distribution of the polypeptide in the former domain. The polypeptide analogous to a photosynthetic bacterial light-harvesting polypeptide binds a pigment, zinc-substituted bacteriochlorophyll a ([Zn]-BChl a), to form the light-harvesting polypeptide/pigment complex. The complex was incorporated into an incompatible binary lipid bilayer combination either the disulfide-based phospholipids bearing oleoyl chains (DO-DO)/1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) or 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC)/DSPC. The lipid-domain-selective organization was confirmed by fluorescence resonance energy transfer (FRET) from a rhodamine-labeled lipid (Lissamine rhodamine B 1,2-diacyl-sn-glycero-3-phosphoethanolamine: either N-Rh-DOPE or N-Rh-DSPE) to the assembled [Zn]-BChl a. Considering the relative affinity of these fluorescent lipid probes, N-Rh-DOPE and N-Rh-DSPE prefer to fluid and gel domains, respectively, because of the similarity of their acyl chains. When the fluorescent lipid probe, N-Rh-DOPE, was applied to the binary lipid system, DO-DO/DSPC, the higher FRET efficiency was observed compared to the counterpart FRET assay using N-Rh-DSPE. In contrast, for the DOPC/DSPC bilayer, the difference in the FRET efficiency between the fluorescent lipid probe systems was negligible. These results suggest that the polypeptide/pigment complex is preferentially incorporated into the disulfide-based lipid domain. Total internal reflection fluorescence (TIRF) microscopic observation of the assembly also evidenced heterogeneous structure by detection of fluorescence from [Zn]-BChl a. [DOI: 10.1380/ejssnt.2005.145]