Fluorescent temporin B derivative and its binding to liposomes.
Temporins are short (10-13 amino acids) and linear antimicrobial peptides first isolated from the skin of the European red frog, Rana temporaria, and are effective against Gram-positive bacteria and Candida albicans. Similarly to other antimicrobial peptides, the association of temporins to lipid membranes has been concluded to underlie their antimicrobial effects. Accordingly, a detailed understanding of their interactions with phospholipids is needed. We conjugated a fluorophore (Texas Red) to a Cys containing derivative of temporin B (temB) and investigated its binding to liposomes by fluorescence spectroscopy. Circular dichroic spectra for the Cys-mutant recorded in the absence and in the presence of phospholipids were essentially similar to those for temB. A blue shift in the emission spectra and diminished quenching by ferrocyanide (FCN) of Texas Red labeled temporin B (TRC-temB) were seen in the presence of liposomes. Both of these changes can be attributed to the insertion of the Texas Red into the hydrophobic region of the bilayer. Resonance energy transfer, steady state anisotropy, and fluorescence lifetimes further demonstrate the interaction of TRC-temB with liposomes to be enhanced by negatively charged phospholipids. Instead, cholesterol attenuates the association of TRC-temB with membranes. The interactions between TRC-temB and liposomes of varying negative surface charge are driven by electrostatics as well as hydrophobicity. Similarly to native temporin B also TRC-temB forms amyloid type fibers in the presence of negatively charged liposomes. This property is likely to relate to the cytotoxic activity of this peptide.
Sood
Rohit
R
Helsinki Biophysics and Biomembrane Group, Medical Biochemistry, Institute of Biomedicine, University of Helsinki, PO Box 63 (Haartmaninkatu 8), Helsinki, FIN-00014 Finland.
Domanov
Yegor
Y
Kinnunen
Paavo K J
PK
D000682:Amyloid / Q000737:chemistry; D000818:Animals; D023181:Antimicrobial Cationic Peptides; D002942:Circular Dichroism; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D014966:Xanthenes / Q000737:chemistry
D000682:Amyloid; D023181:Antimicrobial Cationic Peptides; D005456:Fluorescent Dyes; D008081:Liposomes; D011506:Proteins; D014966:Xanthenes; C104501:temporin; C034657:Texas red