title: A single tryptic fragment of colicin E1 can form an ion channel: stoichiometry confirms kinetics.
abstract: The molecularity of the ion channel formed by peptide fragments of colicin has taken on particular significance since the length of the active peptide has been shown to be less than 90 amino acids and the lumen size at least 8 A. Cell survival experiments show that killing by colicin obeys single-hit statistics, and ion leakage rates from phospholipid vesicles are first order in colicin concentration. However, interpretation in molecular terms is generally complicated by the requirement of large numbers of colicin molecules per cell or vesicle. We have measured the discharge of potential across membranes of small phospholipid vesicles by following the changes in binding of potential sensitive spin labeled phosphonium ions as a function of the number of colicin fragments added. Because of the sensitivity of the method, it was possible to reliably investigate the effect of colicin in a range where there was no more than 0.2 colicins per vesicle. The quantitative results of these experiments yield a direct molecular stoichiometry and demonstrate that one C-terminal fragment of the colicin molecule per one vesicle is sufficient to induce a rapid ion flux in these vesicles. In addition, the experiments confirm earlier findings that the colicin fragments do not migrate from one vesicle to another at pH 4.5. Similar results are obtained with large unilamellar vesicles.
authors.affiliation: Department of Biological Sciences, Columbia University, New York, New York 10027.
authors.forename: F
authors.initials: F
authors.lastname: Levinthal
authors.forename: A P
authors.initials: AP
authors.lastname: Todd
authors.forename: W L
authors.initials: WL
authors.lastname: Hubbell
authors.forename: C
authors.initials: C
authors.lastname: Levinthal
mesh_terms: D003087:Colicins / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D007473:Ion Channels; D008564:Membrane Potentials; D010446:Peptide Fragments / Q000494:pharmacology; D010743:Phospholipids / Q000378:metabolism; D014357:Trypsin
chemical_list: D003087:Colicins; D007473:Ion Channels; D010446:Peptide Fragments; D010743:Phospholipids; D014357:Trypsin