FUNCTIONAL IMMOBILIZATION OF A DNA-BINDING PROTEIN AT A MEMBRANE INTERFACE VIA HISTIDINE TAG AND SYNTHETIC CHELATOR LIPIDS

The coupling of a DNA-binding protein to self-organized lipid monolaye rs is examined at the air-water interface by means of film balance tec hniques and epifluorescence microscopy, We used two recombinant specie s of the heat shock factor HSF24 which differ only in a carboxy-termin al histidine tag that interacts specifically with the nickel-chelating head group of a synthetic chelator lipid, As key function, HSF24 bind s to DNA that contains heat-shock responsible promoter elements. In so lution, DNA-protein complex formation is demonstrated for the wild typ e and fusion protein. Substantial questions of these studies are wheth er protein function is affected after adsorption to lipid layers and w hether a specific docking via histidine tag to the chelator lipid lead s to functional immobilization. Using lipid mixtures that allow a late ral organization of chelator lipids within the lipid film, specific bi nding and unspecific adsorption can be distinguished by pattern format ion of DNA-protein complexes. At the lipid interface, functional DNA-p rotein complexes are only detected, when the histidine-tagged protein was immobilized specifically to a chelator lipid containing monolayer, These results demonstrate that the immobilization of histidine-tag,oe d biomolecules to membranes via chelator lipids is a promising approac h to achieve a highly defined deposition of these molecules at an inte rface maintaining their function.