DeFrND: detergent-free reconstitution into native nanodiscs with designer membrane scaffold peptides

Qian Ren, Jing Wang, Vinay Idikuda, Shanwen Zhang, Jeehae Shin, W. Grant Ludlam, Luis M. Real Hernandez, Sara Zdancewicz, Alex J. B. Kreutzberger, Hucheng Chang, Volker Kiessling, Lukas K. Tamm, Ahmad Jomaa, Ilya Levental, Kirill Martemyanov, Baron Chanda () and Huan Bao ()
Additional contact information
Qian Ren: UF Scripps Biomedical Research
Jing Wang: New York Structural Biology Center
Vinay Idikuda: Washington University School of Medicine
Shanwen Zhang: UF Scripps Biomedical Research
Jeehae Shin: UF Scripps Biomedical Research
W. Grant Ludlam: UF Scripps Biomedical Research
Luis M. Real Hernandez: University of Virginia
Sara Zdancewicz: University of Virginia
Alex J. B. Kreutzberger: University of Virginia
Hucheng Chang: University of Virginia
Volker Kiessling: University of Virginia
Lukas K. Tamm: University of Virginia
Ahmad Jomaa: University of Virginia
Ilya Levental: University of Virginia
Kirill Martemyanov: UF Scripps Biomedical Research
Baron Chanda: Washington University School of Medicine
Huan Bao: UF Scripps Biomedical Research

Nature Communications, 2025, vol. 16, issue 1, 1-16

Abstract: Abstract Membrane scaffold protein-based nanodiscs have facilitated unprecedented structural and biophysical analysis of membrane proteins in a near-native lipid environment. However, successful reconstitution of membrane proteins in nanodiscs requires prior solubilization and purification in detergents, which may impact their physiological structure and function. Furthermore, the detergent-mediated reconstitution of nanodiscs is unlikely to recapitulate the precise composition or asymmetry of native membranes. To circumvent this fundamental limitation of traditional nanodisc technology, we herein describe the development of membrane-solubilizing peptides to directly extract membrane proteins from native cell membranes into nanoscale discoids. By systematically protein engineering and screening, we create a class of chemically modified Apolipoprotein-A1 mimetic peptides to enable the formation of detergent-free nanodiscs with high efficiency. Nanodiscs generated with these engineered membrane scaffold peptides are suitable for obtaining high-resolution structures using single-particle cryo-EM with native lipids. To further highlight the versatility of our approach, we directly extract a sampling of membrane signaling proteins with their surrounding native membranes for biochemical and biophysical interrogations.

Date: 2025
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DOI: 10.1038/s41467-025-63275-8

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