Liquid chromatography MALDI MS/MS for membrane proteome analysis.
Membrane proteins play critical roles in many biological functions and are often the molecular targets for drug discovery. However, their analysis presents a special challenge largely due to their highly hydrophobic nature. We present a surfactant-aided shotgun proteomics approach for membrane proteome analysis. In this approach, membrane proteins were solubilized and digested in the presence of SDS followed by newly developed auto-offline liquid chromatography/matrix-assisted laser desorption ionization (LC/MALDI) tandem MS analysis. Because of high tolerance of MALDI to SDS, one-dimensional (1D) LC separation can be combined with MALDI for direct analysis of protein digests containing SDS, without the need for extensive sample cleanup. In addition, the heated droplet interface used in LC/MALDI can work with high flow LC separations, allowing a relatively large amount of protein digest to be used for 1D LC/MALDI which facilitates the detection of low abundance proteins. The proteome identification results obtained by LC/MALDI are compared to the gel electrophoresis/MS method as well as the shotgun proteomics method using 2D LC/electrospray ionization MS. It is demonstrated that, while LC/MALDI provides more extensive proteome coverage compared to the other two methods, these three methods are complementary to each other and a combination of these methods should provide a more comprehensive membrane proteome analysis.
Zhang
Nan
N
Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2G2.
Li
Nan
N
Li
Liang
L
D002851:Chromatography, High Pressure Liquid; D019073:HT29 Cells; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis*; D020543:Proteome / Q000737:chemistry*; D040901:Proteomics / Q000295:instrumentation / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D008565:Membrane Proteins; D020543:Proteome