Cryo-EM structure of a protein nanopore reveals lipid acyl chains as integral components

Nanopore-based technologies are revolutionizing nucleic acid sequencing and molecular sensing, yet the structural basis by which pore-forming proteins interact with lipid membranes remains poorly defined. The mycobacterial porin MspA is a benchmark biological nanopore owing to its exceptional stability and uniquely narrow constriction, but accurate structural information is limited to protein-only models. Here, we present a 2.5-angstrom resolution cryo-electron microscopy structure of the M2 mutant of MspA embedded in a phospholipid nanodisc, providing a high-resolution experimental observation of MspA in a lipid environment. The structure reveals an annular arrangement of extended lipid acyl chains surrounding the MspA oligomer and contacting the transmembrane barrel. Their resolved densities indicate a substantially more ordered annular lipid environment than would be expected from a largely heterogeneous and nonspecific protein-lipid interface. The structure also demonstrates how specific M2 MspA mutations modulate pore polarity, influencing ion selectivity and transport. By determining the membrane-embedded structure of MspA and revealing an ordered arrangement of acyl chains forming extensive hydrophobic interactions with the protein, this study demonstrates the importance of the surrounding lipid environment in the design of protein nanopores.