Structure-function relationships for a voltage-dependent ion channel: properties of COOH-terminal fragments of colicin E1.

The effects on planar lipid bilayer membranes of carboxyl-terminal fragments derived from the bacteriocin colicin E1 by either proteolysis or CNBr cleavage are indistinguishable from those of the voltage-dependent parent colicin molecule. An upper limit to the length of the COOH-terminal peptide required for channel formation is 152 amino acid residues from the COOH-terminal end, as indicated by the CNBr fragment. In addition, use of carboxypeptidase shows that the COOH-terminal end of the molecule remains on the side of the membrane to which it was added. COOH-terminal peptides of colicin E1 spontaneously associate with oil or hexane droplets in an aqueous system and remain at the interface between the two phases to a significantly greater degree than other colicin E1 fragments or cytochrome c. These results, together with the amino acid sequence, suggest a model wherein the colicin E1 channel is formed first by spontaneous attachment to a membrane of an alpha-helical hairpin centered at a 35-residue hydrophobic region near the COOH-terminal end. Application of a potential of the correct polarity then facilitates a major conformational change in the protein, allowing insertion of the remainder of the COOH-terminal end to form the open channel.