Capillary Zone Electrophoresis-Electrospray Ionization-Tandem Mass Spectrometry as an Alternative Proteomics Platform to Ultraperformance Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry for Samples of Intermediate Complexity

Abstract

We demonstrate the use of capillary zone electrophoresis with an electrokinetically pumped sheath-flow electrospray interface for the analysis of a tryptic digest of a sample of intermediate protein complexity, the secreted protein fraction of Mycobacterium marinum. For electrophoretic analysis, 11 fractions were generated from the sample using reversed phase liquid chromatography; each fraction was analyzed by CZE-ESI-MS/MS, and 334 peptides corresponding to 140 proteins were identified in 165 min of mass spectrometer time at 95% confidence (FDR<0.15%). In comparison, 388 peptides corresponding to 134 proteins were identified in 180 min of mass spectrometer time by triplicate UPLC-ESI-MS/MS analysis each using 250 ng of the unfractionated peptide mixture at 95% confidence (FDR<0.15%). 62% of peptides identified in CZE-ESI-MS/MS and 67% in UPLC-ESI-MS/MS were unique. CZE-ESI-MS/MS favored basic and hydrophilic peptides with low molecular mass. Combining the two data sets increased the number of unique peptides by 53%. Our approach identified more than twice as many proteins as the previous record for CE proteome analysis. CE-ESI-MS/MS is a useful tool for the analysis of proteome samples of intermediate complexity.

Comparing and Combining Capillary Electrophoresis Electrospray Ionization Mass Spectrometry and Nano–Liquid Chromatography Electrospray Ionization Mass Spectrometry for the Characterization of Post-translationally Modified Histones

“…Similar complementary peptide identification was found in recent CE and nano-LC studies by our group and other researchers (32,39,58). CE shows a clear bias toward basic, hydrophilic peptides of low molecular mass.…”

Homeostasis of N-α-Terminal Acetylation of EsxA Correlates with Virulence in Mycobacterium marinum

“…S6 in the supplemental material) (24,58). We measured the levels N-␣-terminally acetylated and unacetylated N-terminal tryptic peptides of EspF and EspB.…”

Developments in Interfacing Designs for CE–MS: Towards Enabling Tools for Proteomics and Metabolomics

“…In a number of publications, Wojcik et al [22] propagate the use of a co-axially, EOF-driven sheath liquid with a borosilicate spray tube emitter for CE-MS [48][49][50][51][52] As capillaries from borosilicate glass are known to be less robust than fused-silica, modest reliability has been reported (personal communication). On the other hand, by using 10 µm i.d.…”