Perfringolysin O association with ordered lipid domains: implications for transmembrane protein raft affinity

Area of Science:

• Membrane biophysics
• Protein-lipid interactions
• Lipid raft dynamics

Background:

• Perfringolysin O (PFO) forms transmembrane β-barrels upon cholesterol interaction.
• PFO is hypothesized to interact with liquid-ordered (LO) lipid domains, known as lipid rafts.
• The molecular basis of transmembrane protein affinity for lipid rafts remains incompletely understood.

Purpose of the Study:

• To investigate the raft affinity of PFO in model membranes with coexisting ordered and disordered lipid domains.
• To elucidate the role of cholesterol and PFO's conformational state in its lipid domain localization.

Main Methods:

• Utilized fluorescence resonance energy transfer (FRET) to quantify PFO's association with lipid domains.
• Employed microscopy on giant unilamellar vesicles (GUVs) to visualize PFO localization.
• Investigated the effect of ceramide on PFO-raft interaction.

Main Results:

• PFO demonstrated intermediate raft affinity, between LW peptide and cholera toxin B.
• PFO's association with ordered domains was not disrupted by ceramide, suggesting shielded cholesterol binding.
• The transmembrane (TM) form of PFO showed lower affinity for ordered domains compared to its prepore form, localizing to domain edges.

Conclusions:

• PFO's TM structure is unfavorable for packing within highly ordered lipid environments.
• Raft localization of TM proteins is controlled by a combination of binding interactions and structural constraints.
• Cellular raft-associated TM proteins may reside in disordered shells within ordered domains to accommodate these factors.