Anal. Chem. 2006, 78, 7309-7316
Analysis of Complex Protein Mixtures with Improved Sequence Coverage Using (CE-MS/MS)n Selynda Garza and Mehdi Moini*
Department of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712
Identification of proteins, in a complex protein mixture, using one-dimensional high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) analysis of its digest, usually suffers from low sequence coverage. There are several reasons for the low coverage including undersampling, wide concentration dynamic range of the proteins in a complex protein mixture, and wide range of electrospray ionization efficiency of peptides under each mobile-phase composition. To address this low sequence coverage, we introduce a novel technique, (CE-MS/MS)n, which utilizes the most significant advantages of CE-MS/MS, including economy of sample size, fast analysis time, and high separation efficiency, to increase the sequence coverage of a complex protein mixture. Based on these characteristics, (CE-MS/MS)n can be performed in which multiple CE-MS/MS subanalyses (injections followed by analyses) are analyzed and experimental variables are manipulated during each CEMS/MS subanalysis in order to maximize sequence coverage. (CE-MS/MS)n is a practical technique since each CE-MS/MS subanalysis consumes
