Electrochemistry-assisted top-down characterization of disulfide-containing proteins

Yun Zhang, Weidong Cui, Hao Zhang, Howard D. Dewald, Hao Chen

Research output: Contribution to journalArticlepeer-review

60 Scopus citations

Abstract

Covalent disulfide bond linkage in a protein represents an important challenge for mass spectrometry (MS)-based top-down protein structure analysis as it reduces the backbone cleavage efficiency for MS/MS dissociation. This study presents a strategy for solving this critical issue via integrating electrochemistry (EC) online with a top-down MS approach. In this approach, proteins undergo electrolytic reduction in an electrochemical cell to break disulfide bonds and then undergo online ionization into gaseous ions for analysis by electron-capture dissociation (ECD) and collision-induced dissociation (CID). The electrochemical reduction of proteins allows one to remove disulfide bond constraints and also leads to increased charge numbers of the resulting protein ions. As a result, sequence coverage was significantly enhanced, as exemplified by ?-lactoglobulin A (24 vs 75 backbone cleavages before and after electrolytic reduction, respectively) and lysozyme (5 vs 66 backbone cleavages before and after electrolytic reduction, respectively). This methodology is fast and does not need chemical reductants, which would have an important impact in high-throughput proteomics research.

Original languageEnglish (US)
Pages (from-to)3838-3842
Number of pages5
JournalAnalytical Chemistry
Volume84
Issue number8
DOIs
StatePublished - Apr 17 2012
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

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