Using phase-resolved vibrational sum-frequency imaging to probe the impact of head-group functionality on...

The substantial diversity in phospholipids within a plasma membrane, varying in tail length, degree of saturation, and head-group functionality, generates widespread structural heterogeneity. This exists both laterally across the membrane through the spontaneous formation of condensed domains that differ from their surrounding expanded phase in density, composition, and molecular packing order, as well as between its two leaflets, which normally maintain significant compositional asymmetry. Of particular importance is the exposure of phosphatidylserine (PS) lipids which is a marker for important physiological processes e.g. apoptosis. Despite this, the molecular-level alterations to the phase-structure of the membrane that result from PS exposure remain generally unknown. In this work, we utilise recently developed phase-resolved azimuthal-scanned sum-frequency generation (SFG) microscopy to investigate structural changes that occur heterogeneously across model membranes as a result of PS-lipid exposure. Specifically, by probing mixed monolayers of 1,2-dipalmitoylphosphatidylcholine (DPPC) and deuterated 1-palmitoyl-2-oleoylphosphatidylcholine (dPOPC) in both the C-H and C-D stretching regions as well as equivalent films with DPPC exchanged with DPPS, we analyse the variations in the apparent phase distributions and domain morphologies, and quantitatively extract the density, composition, and relative out-of-plane packing order for both mixtures. We find that, in these mixtures, DPPS shows vast differences in the domain growth and coalescence behaviour compared to DPPC, as well as in the relative compositions and molecular ordering within each phase. This demonstrates the critical role the head-group plays in the heterogeneous phase structure of the membrane and may give insights into their impact on important physiological processes.