title: Molecular characterization of the interaction of crotamine-derived nucleolar targeting peptides with lipid membranes.
abstract: A novel class of cell-penetrating, nucleolar-targeting peptides (NrTPs), was recently developed from the rattlesnake venom toxin crotamine. Based on the intrinsic fluorescence of tyrosine or tryptophan residues, the partition of NrTPs and crotamine to membranes with variable lipid compositions was studied. Partition coefficient values (in the 10(2)-10(5) range) followed essentially the compositional trend POPC:POPG≤POPG<POPC≤POPC:cholesterol. Leakage assays showed that NrTPs induce minimal lipid vesicle disruption. Fluorescence quenching of NrTPs, either by acrylamide or lipophilic probes, revealed that NrTPs are buried in the lipid bilayer only for negatively-charged membranes. Adoption of partial secondary structure by the NrTPs upon interaction with POPC and POPG vesicles was demonstrated by circular dichroism. Translocation studies were conducted using a novel methodology, based on the confocal microscopy imaging of giant multilamellar vesicles or giant multivesicular liposomes. With this new procedure, which can now be used to evaluate the membrane translocation ability of other molecules, it was demonstrated that NrTPs are able to cross lipid membranes even in the absence of a receptor or transmembrane gradient. Altogether, these results indicate that NrTPs interact with lipid bilayers and can penetrate cells via different entry mechanisms, reinforcing the applicability of this class of peptide as therapeutic tools for the delivery of molecular cargoes.
authors: Instituto de Medicina Molecular, Faculdade de Medicina da Universidade de Lisboa, Lisbon, Portugal.
authors: Margarida
authors: M
authors: Rodrigues
authors: Andrea
authors: A
authors: Santos
authors: Beatriz G
authors: BG
authors: de la Torre
authors: Gandhi
authors: G
authors: Rádis-Baptista
authors: David
authors: D
authors: Andreu
authors: Nuno C
authors: NC
authors: Santos
mesh_terms: D000595:Amino Acid Sequence; D002466:Cell Nucleolus / Q000378:metabolism*; D002942:Circular Dichroism; D003435:Crotalid Venoms / Q000378:metabolism*; D005453:Fluorescence; D008563:Membrane Lipids / Q000378:metabolism*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D003435:Crotalid Venoms; D008563:Membrane Lipids; D010455:Peptides; C010563:crotamine