A study of the membrane-bound topology of the colicin E1 channel domain
Colicin E1 is a member of the channel-forming subfamily of colicins. Despite the 2.5 Å resolution X-ray structure of the soluble colicin E1 channel domain (P190) (Elkins 'et al., Structure,' 5: 443-458, 2001), the membrane-bound structure of P190 peptide is poorly understood. In this thesis, a study of the membrane-bound topology of helix III of P190 by site-directed fluorescence labelling was undertaken. It was discovered that the amphipathic helix III retains its alpha helical nature upon membrane association. Interestingly, the helix adopted a tilted and extended orientation in the membrane-bound state. The overall membrane topology of P190 was also studied using fluorescence resonance energy transfer (FRET). This study used nine intra-molecular distances to elucidate the conformational changes occurring within P190 at the membrane surface. The data were used to generate a low resolution three-dimensional model that provided new insights into the disposition of the helices of P190 upon membrane-binding.