Nanodiscs can preferentially enrich transmembrane proteins in a bottom-up proteomic analysis workflow
Year: 2026
Type: preprint
Abstract: Membrane proteins are critically important pharmaceutical targets and thus their analytical characterization is vital to drug development efforts. However, compared to cytosolic proteins, membrane proteins are generally more difficult to purify and/or analytically characterize in part due to their amphipathic nature. Nanodiscs (NDs) have become a popular membrane mimetic system offering a well-defined lipid bilayer environment to stabilize membrane proteins for in vitro analyses. NDs based on membrane scaffold proteins (MSPs) offer advantages in terms of their amenability to purification and control over lipid composition. Here we investigated the utility of MSP NDs having different diameters and lipid compositions to enrich membrane proteins isolated from cells for bottom-up proteomic characterization. Specifically, we coupled a robust ND formation and purification workflow with liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to determine the identity and relative abundance of proteins incorporated into NDs made using different synthesis conditions. We found that the ND diameter had an effect on the relative enrichment of detected proteins and protein classes; however, lipid composition did not have a significant effect. The data indicated that this ND-based approach selectively allowed for the detection of membrane proteins and membrane associated proteins over cytosolic protein contaminants. Furthermore, we found that transmembrane proteins, which are particularly hard to extract and analyze using traditional membrane mimetics, were significantly enriched when using NDs compared to detergent micelles. These results suggest that MSP ND-based approaches could be applied to the selective enrichment of membrane proteins from cells in a manner that could be enabling for both proteomic and other analytical characterization approaches.
Source: ChemRxiv
Institution: University of Michigan
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7214101636