## 6 Lipids in SMALPs  
The design of SMA as an effective mild solubilizer of virtually any lipid bilayer or memtein relies on the short styrene sidechains which insert into hydrophobic material [20]. The specific lipid species bound to proteins in SMALPs can most easily be identified using MS. This was demonstrated for SMALP'd insect cell membranes containing the equilibrative nucleoside transporter-1 (hENT1). This protein solubilizes with 16 PC and 2 PE molecules but no polyunsaturated lipids based on electrospray ionization (ESI) MS data [58]. To prevent protein degradation and inactivation, the XIRAN 30010 polymer had been added at a relatively low concentration (0.25% \(w/v\)) along with cholesteryl hemisuccinate at low temperature. The resulting protein in nanodiscs shows the predicted inhibitor binding activity, in contrast to protein solubilized in decyl maltoside.  
The phospholipids which co-extract with three bacterial membrane proteins can be determined using LC-MS/MS methods [59]. SMA copolymer does not preferentially solubilize any specific types of phospholipids from the membrane, and thus is suitable for unbiased detection of protein-bound phospholipids. The ZipA and PgpB proteins contain 1 and 6 transmembrane helices, respectively, and bind similar ratios of various phospholipids, with higher levels of monounsaturated PE and PG species. In contrast the membrane associated protein FtsA bound preferentially to a distinct profile of PE and PG lipids, presumably as it only inserts part of the way into the inner leaflet of the cytoplasmic membrane.  
The GlpG rhomboid proteases from _Escherichia coli_ and _Vibrio cholerae_ have been solubilized using DIBMA, SMA(3:1), and SMA(2:3:1), and their yields and activities compared [60]. Native-like protease activities and high stabilities were found after purification with SMA(3:1) and DIBMA but not with conventional detergents. GlpG binds a variable number of about 50 phospholipids of different chain lengths and degrees of saturation depending on the cell type and temperature based on electrospray ionization and collision-induced dissociation MS data [61].  
While the polydispersity of synthetic polymers such as SMA allows a large range of lipids and proteins to be solubilized effectively, this also poses technical issues for MS analysis of SMALPs and multimeric proteins. In particular, the broad range of polymer masses, stoichiometries, and nanodisc diameters can lead to a peak broadening of the assemblies when analyzed using MS. Moveover, the voltages in ESI-MS experiments may be insufficient to release the contained protein from strong interactions with lipid sheets inside the SMALP. This is being addressed by laser-induced liquid bead ion desorption (LILBID) MS, which allows nanodiscs to be transferred within 50 \(\upmu\)m diameter droplets that experience soft or hard laser pulses. This results in detection of memteins that can be progressively dissociated in their component lipid-bound subunits, as demonstrated by the GlpG monomers, KtrB potassium channel dimers, and AcrB trimers [62]. This approach allows novel multimeric states to be resolved, including the monomeric and dimeric states of the sodium-solute symporter and KimA proteins, suggesting wider utility.