Home is where the lipids are: a comparison of MSP and DDDG nanodiscs for membrane protein research

Nanodiscs have emerged as a powerful tool for studying membrane proteins in a lipid bilayer, with the standard approach relying on MSP-based nanodiscs that use detergent-mediated lipid exchange and encapsulation by MSP rings. However, this method may introduce artefacts from MSP interactions with the target protein and the nanodiscs constrained size. Here, we compare MSP-based nanodiscs with an alternative system using the amphiphile dodecyl-diglucoside (DDDG), which directly extracts membrane proteins along with their surrounding lipids from the cell membrane. Using a glutamate transporter homolog (GltTk) from <i>Thermococcus kodakarensis</i> as a model, we assessed the efficiency of extraction and purification, thermal stability, and substrate binding capacity of GltTk in each of the two nanodisc systems. Our findings demonstrate that DDDG-based nanodiscs are comparable to MSP-based nanodiscs but may provide greater conformational flexibility and avoid possible artefacts due to MSP-GltTk interactions. Consequently, they provide a competent alternative to MSP-based nanodiscs through direct extraction, thereby preserving the proteins native lipid environment. Both approaches support structural and functional studies, but their suitability depends on the specific application. MSP-based nanodiscs remain advantageous for studies requiring well-defined lipid compositions, while DDDG nanodiscs offer distinct advantages for investigating proteins where native lipids and conformational freedom are critical.