Amphipathic Helices in Antimicrobial Peptides

The amphipathic α-helix is a fundamental structural motif in antimicrobial peptides, enabling selective membrane disruption.

What Is an Amphipathic Helix?

Definition An α-helix with: - **Hydrophobic face** — Nonpolar residues (Leu, Ile, Phe, Trp) - **Hydrophilic face** — Polar/charged residues (Lys, Arg, Ser) - **Segregation** — Faces on opposite sides of the helix

The Hydrophobic Moment Quantitative measure of amphipathicity: - Vector sum of residue hydrophobicities - Higher = more amphipathic - Optimal range for AMP activity

LL-37: The Human Cathelicidin

Structure - 37 amino acids (LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES) - Forms amphipathic α-helix - Disordered in aqueous solution - Folds upon membrane contact

Amphipathic Properties - Hydrophobic face: Leu, Phe, Ile, Val residues - Cationic face: Lys, Arg residues (+6 net charge) - Clear segregation visualized by helical wheel

Membrane Selectivity

Why AMPs Target Bacteria

The Selectivity Mechanism 1. Cationic peptide attracted to anionic bacterial surface 2. Hydrophobic face inserts into lipid bilayer 3. Cholesterol in mammalian membranes prevents deep insertion 4. Bacterial membrane disrupted; mammalian cells spared

Models of Membrane Disruption

Barrel-Stave Model - Peptides form transmembrane pores - Hydrophobic faces contact lipid tails - Hydrophilic faces line aqueous pore - Requires defined stoichiometry

Toroidal Pore Model - Peptides cause lipid headgroups to bend inward - Continuous surface of peptide + lipid - Lower peptide:lipid ratio than barrel-stave - **Most common for α-helical AMPs**

Carpet Model - Peptides accumulate on membrane surface - Cover like a "carpet" - At threshold concentration, membrane disintegrates - Detergent-like mechanism

Aggregate Channel Model - Disordered peptide aggregates in membrane - Transient, heterogeneous pores - Less organized than other models

Structure-Activity Relationships

Optimization Parameters | Parameter | Effect of Increase | |-----------|-------------------| | Hydrophobicity | ↑ Activity, ↑ Toxicity | | Net charge | ↑ Selectivity (to a point) | | Helix stability | ↑ Activity, ↑ Resistance | | Amphipathicity | ↑ Activity (optimal range) |

The Toxicity Trade-off - Very hydrophobic peptides insert into any membrane - Must balance potency against host toxicity - Typically +2 to +9 net charge optimal

Induced Folding

Disorder-to-Order Transition Many AMPs are unstructured in solution: - Avoids self-association - Reduces toxicity during transport - Fold triggers activity

Membrane-Induced Folding 1. Initial electrostatic attraction 2. Partitioning into membrane interface 3. Helix formation in lipid environment 4. Insertion and disruption

Implications for Design - Can engineer "conditional" activity - Stability in solution may not predict activity - Membrane environment is the true context

Therapeutic Engineering

Improving Selectivity - Tune charge/hydrophobicity balance - Use D-amino acids for protease resistance - Cyclization for stability

Reducing Hemolysis - Proline substitutions to break helix - Charge modifications - PEGylation

Example: Pexiganan (MSI-78) - Synthetic magainin analog - 22 amino acids, amphipathic helix - Reached Phase 3 for diabetic foot ulcers - Improved potency and stability over natural peptide