title: Decoding distinct membrane interactions of HIV-1 fusion inhibitors using a combined atomic force and fluorescence microscopy approach.
abstract: Enfuvirtide and T-1249 are two potent HIV-1 fusion inhibitor peptides. Recent studies indicate that lipids play an important role in the mode of action of those bioactive molecules. Using a combined tandem atomic force microscopy (AFM)-epifluorescence microscopy approach, we studied the interaction of both enfuvirtide and T-1249 with supported lipid bilayers. Fluid (ld)-gel (so) and ld-liquid ordered (lo) phase-separated membrane systems were tested. Results, especially for T-1249, show significant lipid membrane activity at a 15μM peptide concentration. T-1249, in opposition to enfuvirtide, induces an increase in membrane surface roughness, decrease in membrane fluidity, bilayer thinning at ld domains and disruption of the so domain borders. In terms of structural properties, both enfuvirtide and T-1249 possess distinct functional hydrophobic and amphipathic domains of HIV gp41. While enfuvirtide only yields the tryptophan-rich domain (TRD), T-1249 possesses both TRD and pocket-binding domain (PBD). TRD increases the hydrophobicity of the peptide while PBD enhances the amphipathic characteristics. As such, the enhanced membrane activity of T-1249 may be explained by a synergism between its amphipathic N-terminal segment and its hydrophophic C-terminal. Our findings provide valuable insights on the molecular-level mode of action of HIV-1 fusion inhibitors, unraveling the correlation between their structural properties and membrane interactions as a factor influencing their antiviral activity. Ultimately, this work validates the applicability of a combined AFM and fluorescence approach to evaluate the mechanic and structural properties of supported lipid bilayers upon interaction with membrane-active peptides.
authors.affiliation: Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.
authors.forename: Henri G
authors.initials: HG
authors.lastname: Franquelim
authors.forename: Diana
authors.initials: D
authors.lastname: Gaspar
authors.forename: A Salomé
authors.initials: AS
authors.lastname: Veiga
authors.forename: Nuno C
authors.initials: NC
authors.lastname: Santos
authors.forename: Miguel A R B
authors.initials: MA
authors.lastname: Castanho
mesh_terms: D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D000077560:Enfuvirtide; D015700:HIV Envelope Protein gp41 / Q000494:pharmacology*; D023581:HIV Fusion Inhibitors / Q000494:pharmacology*; D015658:HIV Infections / Q000188:drug therapy / Q000378:metabolism / Q000821:virology; D015497:HIV-1 / Q000187:drug effects*; D006801:Humans; D008051:Lipid Bilayers / Q000378:metabolism*; D008560:Membrane Fluidity; D008563:Membrane Lipids / Q000378:metabolism; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence; D010446:Peptide Fragments / Q000494:pharmacology*
chemical_list: D015700:HIV Envelope Protein gp41; D023581:HIV Fusion Inhibitors; D008051:Lipid Bilayers; D008563:Membrane Lipids; D010446:Peptide Fragments; D000077560:Enfuvirtide