Hey everyone! Would be really appreciated for advice. Trying to solubilize plasma membrane with Styrene - maleic acid (SMA) copolymers. Following all the instructions making a 5% SMA solution, looks like nothing special. But after incubation overnight with cell membrane turns out (via WB) neither nanodiscs are created. Did the same with DDM detergent and it worked well! So if anyone has worked with SMALP nanodiscs I would be so happy for recommendations and advice:)
Solubilization with SMA copolymers can be somewhat different than detergent, for example polymer solubilization is normally done around room temperature (or even higher, compared to on ice for detergent). Generally you can expect the copolymers to have poorer extraction efficiency compared to harsh detergents (they make up for it in terms of stability and being in the native environment). The exact SMA copolymer you are using can have a big influence, generally the 2:1 (ratio styrene-to-maleic acid, or otherwise 3:1) is recommended. Other factors like ionic strength, pH and divalent cations can also play a role. Please see the following publication for more details Article Factors influencing the solubilization of membrane proteins ...
, of course that study was done on E.coli but there will be similarities with other membrane systems. Is your final SMA concentration 2.5% then? Although this is the standard starting recommendation it is quite high and you may find better results with a lower concentration (e.g. 0.5% - 1%), especially when it comes to protein purification. Perhaps solubilization was successful but the polymer is interfering with the antibody binding in WB?I hope that this helps :) Good luck with your experiments!
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