Trends in the use of amphipathic environments and future perspectives for determining the structure of membrane proteins by cryo-EM.

Membrane proteins (MPs) play essential roles in a wide range of cellular processes and represent major therapeutic targets. Nevertheless, their structural and functional characterization remains challenging due to inherent difficulties in production, extraction, and stabilization outside their native lipid environment. The rise of cryogenic electron microscopy (cryo-EM) has markedly accelerated the structure determination of MPs through single-particle analysis (SPA). A systematic examination of high-resolution cryo-EM SPA structures deposited in the Protein Data Bank (PDB) over the past two years provides a comprehensive overview of the most frequently used amphipathic environments. We discuss the strengths and limitations of each approach and underscore the ongoing need to develop near-native environment strategies to improve the interpretation of MP structures under membrane-like conditions.