Surfactant-Mediated Structural Modulations to Planar, Amphiphilic Multilamellar Stacks

Abstract

The hydrophobic effect, a ubiquitous process in biology, is a primary thermodynamic driver of amphiphilic self-assembly. It leads to the formation of unique morphologies including two highly important classes of lamellar and micellar mesophases. The interactions between these two types of structures and their involved components have garnered significant interest because of their importance in key biochemical technologies related to the isolation, purification, and reconstitution of membrane proteins. This work investigates the structural organization of mixtures of the lamellar-forming phospholipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and two zwitterionic micelle-forming surfactants, being n-dodecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (Zwittergent 3-12 or DDAPS) and 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (O-Lyso-PC), when assembled by water vapor hydration with X-ray diffraction measurements, brightfield optical microscopy, wide-field fluorescence microscopy, and atomic force microscopy. The results reveal that multilamellar mesophases of these mixtures can be assembled across a wide range of POPC to surfactant (POPC:surfactant) concentration ratios, including ratios far surpassing the classical detergent-saturation limit of POPC bilayers without significant morphological disruptions to the lamellar motif. The mixed mesophases generally decreased in lamellar spacing (D) and headgroup-to-headgroup distance (D_hh) with a higher concentration of the doped surfactant, but trends in water layer thickness (D_w) between each bilayer in the stack are highly variable. Further structural characteristics including mesophase topography, bilayer thickness, and lamellar rupture force were revealed by atomic force microscopy (AFM), exhibiting homogeneous multilamellar stacks with no significant physical differences with changes in the surfactant concentration within the mesophases. Taken together, the outcomes present the assembly of unanticipated and highly unique mixed mesophases with varied structural trends from the involved surfactant and lipidic components. Modulations in their structural properties can be attributed to the surfactant's chemical specificity in relation to POPC, such as the headgroup hydration and the hydrophobic chain tail mismatch. Taken together, our results illustrate how specific chemical complexities of surfactant-lipid interactions can alter the morphologies of mixed mesophases and thereby alter the kinetic pathways by which surfactants dissolve lipid mesophases in bulk aqueous solutions.

Figure 1

Chemical structure of the experimental amphiphiles POPC, DDAPS, and O-Lyso-PC. Chemical structures of (a) 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), (b) n-dodecyl-N,N-dimethyl-3-ammonio-1-propanesulfonate (DDAPS), and (c) 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (O-Lyso-PC).

Figure 2

Experimental XRD data of POPC:DDAPS mesophases. A stacked plot of the intensities of the X-ray diffraction peaks of various POPC:DDAPS multilamellar mesophases was constructed. The molar ratios plotted include 1:0, 1:1, 2:3, 1:2, and 1:4 POPC:DDAPS.

Figure 3

Lamellar structure of POPC:DDAPS multilamellar mesophases by molar fraction. (a) Average electron densities normal to the bilayers were assembled from the diffraction peaks and plotted with an arbitrary scale. The molar ratios plotted include 1:0 (black), 1:1 (red), 2:3 (blue), 1:2 (green), and 1:4 (orange) POPC:DDAPS. (b) Lamellar spacing (D) of POPC:DDAPS multilamellar mesophases was deduced by XRD and plotted by the molar fraction of DDAPS. (c) The headgroup-to-headgroup distances (D_hh) of POPC:DDAPS multilamellar mesophases were calculated by XRD and plotted by molar fraction of DDAPS. (d) Water layer thickness (D_w) was calculated from D and D_hh and was similarly plotted.

Figure 4

AFM topographic images of POPC:DDAPS mesostructures. (a) AFM topographic image of 1:0 molar ratio POPC:DDAPS. (b) A 3D display of (a). (c) The AFM topographic image of the area indicated by the red square in (a). (d) A cursor profile of height over the mesophase’s surface as indicated by the red line in (c). (e) An AFM topographic image of 1:1 molar ratio POPC:DDAPS. (f) A 3D display of (e). (g) The AFM topographic image of the area indicated by the red square in (e). (h) A cursor profile of height over the mesophase’s surface as indicated by the red line in (g). (i) An AFM topographic image of 1:2 molar ratio POPC:DDAPS. (j) A 3D display of (i). (k) The AFM topographic image of the area indicated by the red square in (i). (l) A cursor profile of height over the mesophase’s surface as indicated by the red line in (k). (m) An AFM topographic image of 1:3 molar ratio POPC:DDAPS. (n) A 3D display of (m). (o) The AFM topographic image of the area indicated by the red square in (m). (p) A cursor profile of height over the mesophase’s surface as indicated by the red line in (o). (q) An AFM topographic image of 1:4 molar ratio POPC:DDAPS. (r) A 3D display of (q). (s) The AFM topographic image of the area indicated by the red square in (q). (t) A cursor profile of height over the mesophase’s surface as indicated by the red line in (s). Blue scale bar = 2 μm, white scale bar = 20 μm.

Figure 5

Bilayer thickness and rupture force of POPC:DDAPS multilamellar mesophases by molar fraction as measured by AFM. (a) Bilayer thickness (D_t) of POPC:DDAPS multilamellar mesophases was deduced by AFM across the entire stack, averaged, and plotted by molar fraction of DDAPS. Error bars are standard deviations. (b) Bilayer rupture force (F_r) of POPC:DDAPS multilamellar mesophases was similarly elucidated and plotted by molar fraction of DDAPS. Error bars are standard deviations.

Figure 6

Experimental XRD data of POPC:O-Lyso-PC mesophases. A stacked plot of the intensities of the X-ray diffraction peaks of various POPC:O-Lyso-PC multilamellar mesophases was constructed. The molar ratios plotted include 1:0, 1:1, 2:3, 1:2, and 1:4 POPC:O-Lyso-PC.

Figure 7

Lamellar structure of POPC:O-Lyso-PC multilamellar mesophases by the molar fraction. (a) Average electron densities normal to the bilayers were assembled from the diffraction peaks and plotted with an arbitrary scale. The molar ratios plotted include 1:0 (black), 1:1 (red), 2:3 (blue), 1:2 (green), and 1:4 (orange) POPC:DDAPS. (b) Lamellar spacing (D) of POPC:O-Lyso-PC multilamellar mesophases were deduced by XRD and plotted by molar fraction of O-Lyso-PC. (c) The headgroup-to-headgroup distance (D_hh) of POPC:O-Lyso-PC multilamellar mesophases was calculated by XRD and plotted by molar fraction of O-Lyso-PC. (d) Water layer thickness (D_w) was calculated from D and D_hh and similarly plotted.

Figure 8

Water vapor-mediated assembly of multilamellar mesophases. A cartoon representation of a proposed mechanism of the water vapor hydration of the POPC:surfactant mixtures into multilamellar mesophases. Dried mixtures of POPC (purple cylinders) and surfactants (yellow cones) are hydrated by the surrounding water (blue and red models) within the humidity chamber, and lyotropic networks of amphiphiles and water assemble. In this mechanism, the morphological consequences of differentiated spontaneous curvature (and therefore solubilization) are repressed upon hydration due to energetic considerations of bilayer bending and the hydration network reorganizing.