Lipid acyl chain length and unsaturation modulate membrane surface charge and interactions with amphiphilic DNA nanoprobes

Many cellular pathways rely on the active regulation of membrane composition to fine-tune key physico-chemical properties, including lipid packing, fluidity, and surface charge. While intrinsic membrane charge is often attributed to specific lipid headgroups and membrane-bound proteins, the contribution of acyl-chain chemistry to the electrostatic profile of membrane surfaces remains unexplored. Here, we systematically investigate how variations in acyl chain length and unsaturation modulate the biophysical and electrostatic properties of zwitterionic lipid membranes. Using amphiphilic DNA nanoprobes as model charged biomolecules, we reveal how lipid packing, fluidity, and membrane phase collectively govern surface charge and interactions with nanoprobes, delineating relationships that persist in the presence of anionic lipids. We further demonstrate that the identity and hydrophobicity of membrane anchors in nanoprobes significantly influence binding, providing a means to modulate membrane association through programmable strategies. By establishing design principles that link acyl chain chemistry to surface charge and biomolecular attachment, our findings provide a mechanistic framework to engineer selective membrane interactions. Beyond their direct applicability to biomimetic platforms and synthetic cell engineering, these insights hold broad relevance to lipid-based vaccine nanotechnologies and the fundamental understanding of membrane-biomolecule interactions in living cells.