Using SANS with contrast-matched lipid bicontinuous cubic phases to determine the location of encapsulated peptides, proteins, and other biomolecules

An understanding of the location of peptides, proteins, and other biomolecules within the bicontinuous cubic phase is crucial for understanding and evolving biological and biomedical applications of these hybrid biomolecule lipid materials, including during in meso crystallization and drug delivery. While theoretical modeling has indicated that proteins and additive lipids might phase separate locally and adopt a preferred location in the cubic phase, this has never been experimentally confirmed. We have demonstrated that perfectly contrast-matched cubic phases in D2O can be studied using small-angle neutron scattering by mixing fully deuterated and hydrogenated lipid at an appropriate ratio. The model transmembrane peptide WALP21 showed no preferential location in the membrane of the diamond cubic phase of phytanoyl monoethanolamide and was not incorporated in the gyroid cubic phase. While deuteration had a small effect on the phase behavior of the cubic phase forming lipids, the changes did not significantly affect our results.