SAP(E) - A cell-penetrating polyproline helix at lipid interfaces

Area of Science:

• Biophysics
• Molecular Biology
• Biochemistry

Background:

• Cell-penetrating peptides (CPPs) facilitate cellular uptake of molecules but their internalization mechanisms remain unclear.
• Sweet arrow peptides (SAPs) are negatively charged CPPs adopting a polyproline II helix, distinct from typical positively charged CPPs.
• Proposed SAP internalization involves surface aggregation and endocytosis, yet direct molecular-level observation is challenging.

Purpose of the Study:

• To investigate the molecular interactions between SAP(E) peptides and model cell membranes.
• To elucidate the role of membrane charge and structure in SAP(E) peptide binding and internalization.
• To directly observe peptide-membrane interactions using advanced spectroscopy.

Main Methods:

• Sum Frequency Generation (SFG) vibrational spectroscopy was employed to study SAP(E) interactions.
• Experiments utilized differently charged model membranes in both mono- and bi-layer configurations.
• Spectroscopic analysis focused on peptide structure and binding sites on the membrane.

Main Results:

• SFG spectroscopy revealed structural changes in SAP(E) upon initial binding to model membranes.
• Peptide-membrane interaction strength and mode depend on the lipid headgroup charge.
• Phosphocholine headgroups were identified as favorable binding sites for SAP(E).
• SAPs interact with the membrane surface, specifically the headgroup region, without penetrating the hydrophobic core.

Conclusions:

• SAP(E) binding to model membranes is initiated by surface interactions accompanied by peptide structural alterations.
• The charge of the lipid headgroup significantly influences SAP(E) binding affinity.
• Unlike some CPPs, SAP(E) does not insert into the lipid bilayer's hydrophobic core, suggesting a distinct internalization pathway.