Ionic liquid 1-butyl-3-methyl imidazolium tetrafluoroborate for shotgun membrane proteomics

The solubility and digestion efficiency are two crucial factors that might affect the identification of integral membrane proteins (IMPs). In this work, 1% (v/v) ionic liquid (IL), 1-butyl-3-methyl imidazolium tetrafluoroborate (BMIM BF(4)), added in NH(4)HCO(3) buffer (pH 8.3), was applied as a sample preparation buffer for IMPs analysis. Compared to the commonly used sodium dodecyl sulfate and methanol methods, the number of identified IMPs from rat brain by microcolumn reversed phase liquid chromatography (μRPLC)-electrospray ionization tandem mass spectrometry (ESI-MS/MS) was improved by over three times, which might be due to the fact that BMIM BF(4) offered high solubilizing ability for IMPs and good compatibility for tryptic digestion. Furthermore, compared to Rapigest and urea methods, with BMIM BF(4) method, the number of identified IMPs from rat brain could be improved 25% and 80%, respectively, which might be contributed to the good solubilizing ability and high thermal stability of such IL. With the sample treated by BMIM BF(4) method, by 2D-nanoSCX-RPLC-ESI-MS/MS, 1,450 non-redundant proteins and 7,978 unique peptides were identified from rat brain, and 418 proteins contained at least one predicted transmembrane domain, with false discovery rates of less than 1% for peptide identification, and at least two identified unique peptides per protein. All these results demonstrate that the BMIM BF(4) method is of high potential for the large-scale identification of IMPs.