Structural characterisation of the natural membrane-bound state of melittin: a fluorescence study of a dansylated analogue.
The binding of a dansylated analogue of melittin (DNC-melittin) to natural membranes is described. The cytolytic peptide from honey bee venom melittin was enzymatically labelled in its glutamine-25 with the fluorescent probe monodansylcadaverine using guinea pig liver transglutaminase. The labelled peptide was characterised functionally in cytolytic assays, and spectroscopically by circular dichroism and fluorescence. The behaviour of DNC-melittin was, in all respects, indistinguishable from that of the naturally occurring peptide. We used resonance energy transfer to measure the state of aggregation of melittin on the membrane plane in synthetic and natural lipid bilayers. When bound to erythrocyte ghost membranes, the extent of energy transfer was found to be equivalent to when bound to small unilamellar vesicles of phosphatidylcholine. Our results correlate best with a proposed model in which the initial interaction between melittin and the red blood cells could be merely electrostatic and the peptide remains in a low alpha-helical conformation. The next step would be a peptide stabilisation in the membrane in a monomeric alpha-helical conformation that would imply the collapse of the membrane structure and liberation of the cell contents.
Pérez-Payá
E
E
Departament de Bioquímica i Biologia Molecular, Universitat de València, Burjassot, Spain.
Dufourcq
J
J
Braco
L
L
Abad
C
C
D000818:Animals; D001665:Binding Sites; D002103:Cadaverine / Q000031:analogs & derivatives / Q000494:pharmacology; D002942:Circular Dichroism; D004735:Energy Transfer; D005456:Fluorescent Dyes; D005973:Glutamine; D006168:Guinea Pigs; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D008051:Lipid Bilayers; D008099:Liver / Q000201:enzymology; D008555:Melitten / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D011503:Transglutaminases / Q000378:metabolism
C109189:DNC-melitten; D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D005973:Glutamine; D008555:Melitten; D011503:Transglutaminases; C008542:monodansylcadaverine; D002103:Cadaverine