title: Membrane-binding amphipathic alpha-helical peptide derived from CTP:phosphocholine cytidylyltransferase.
abstract: A peptide corresponding to a portion of the amphipathic alpha-helical region of CTP:phosphocholine cytidylyltransferase was synthesized. This region of the enzyme was proposed to be the membrane-binding domain [Kalmar, G.B., Kay, R.J., Lachance, A., Aebersold, R., & Cornell, R.B. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 6029]. We have shown that the peptide is physically associated with PG vesicles. CD of the peptide in buffer suggested a primarily random structure, while, in the presence of trifluoroethanol, the peptide was alpha-helical. Anionic lipid vesicles promoted an alpha-helical conformation, whereas neutral or cationic lipid vesicles did not alter the random structure of the peptide, suggesting a selective stabilization of the alpha-helix by anionic membranes. The fluorescence of the single tryptophan residue, which lies on the hydrophobic face of the amphipathic alpha-helix, was studied. Anionic lipid vesicles specifically induced a shift in the fluorescence to a lower wavelength. Fluorescence quenching by the aqueous-phase quencher, I-, and the lipid-phase quencher 9,10-dibromo-PC was used to determine the accessibility of the tryptophan to each of these environments. The presence of anionic lipid vesicles, but not nonanionic lipid vesicles, decreased the quenching by I- suggesting that, in the presence of anionic lipids, the tryptophan residue is poorly accessible to the aqueous I-. Dibromo-PC significantly quenched the fluorescence only when present in anionic vesicles, confirming the membrane location of the tryptophan residue and the lipid specificity of this interaction.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Chemistry, Simon Fraser University, Burnaby, British Columbia, Canada.
authors.forename: J E
authors.initials: JE
authors.lastname: Johnson
authors.forename: R B
authors.initials: RB
authors.lastname: Cornell
mesh_terms: D000595:Amino Acid Sequence; D000838:Anions; D001665:Binding Sites; D001966:Bromine; D002462:Cell Membrane / Q000201:enzymology; D019916:Choline-Phosphate Cytidylyltransferase; D002942:Circular Dichroism; D007454:Iodides; D008565:Membrane Proteins / Q000138:chemical synthesis / Q000737:chemistry*; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000737:chemistry*; D010446:Peptide Fragments / Q000138:chemical synthesis*; D010743:Phospholipids / Q000378:metabolism*; D017433:Protein Structure, Secondary; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry
chemical_list: D000838:Anions; D007454:Iodides; D008565:Membrane Proteins; D010446:Peptide Fragments; D010743:Phospholipids; D014364:Tryptophan; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; D001966:Bromine