Imaging Cu²⁺ binding to charged phospholipid membranes by high-throughput second harmonic wide-field microscopy

Area of Science:

• Biophysics
• Membrane Biophysics
• Ion-Membrane Interactions

Background:

• Divalent copper ions (Cu²⁺) are vital for cellular functions like hormone synthesis and energy production.
• Membrane hydration is essential for Cu²⁺ interactions, yet its role remains understudied.
• Understanding Cu²⁺-lipid interactions is key to elucidating cellular signaling and transport.

Purpose of the Study:

• To investigate the role of membrane hydration in Cu²⁺ interactions using advanced microscopy.
• To analyze the impact of Cu²⁺ binding on lipid membrane structure and interfacial properties.
• To quantify differences in membrane potential and binding affinities based on lipid head groups.

Main Methods:

• Utilized high-throughput wide-field second harmonic (SH) microscopy.
• Studied hydrated freestanding Montal-Müller lipid membranes composed of specific phospholipids.
• Introduced Cu²⁺ to one leaflet of symmetric lipid bilayers while maintaining ionic strength.

Main Results:

• Observed transient domains with high SH intensity, indicating Cu²⁺ binding to lipid head groups.
• Demonstrated Cu²⁺-induced charge neutralization on one membrane side, exposing ordered interfacial water.
• Quantified higher interfacial membrane potential in phosphatidic acid domains compared to phosphatidylserine domains.

Conclusions:

• Cu²⁺ binding significantly alters membrane interfacial properties and water organization.
• Lipid head group identity (phosphatidic acid vs. phosphatidylserine) influences Cu²⁺ binding affinity and electrostatic interactions.
• The study provides insights into the dynamic interplay between ions, lipids, and water at the cell membrane interface.