Absolute Quantitation of Proteins by Coulometric Mass Spectrometry

Pengyi Zhao, Qi Wang, Manpreet Kaur, Yong Ick Kim, Howard D. Dewald, Olivier Mozziconacci, Yong Liu, Hao Chen

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Accurate quantification is essential in the fields of proteomics, clinical assay, and biomarker discovery. Popular methods for absolute protein quantitation by mass spectrometry (MS) involve the digestion of target protein and employ isotope-labeled peptide internal standards to quantify chosen surrogate peptides. Although these methods have gained success, syntheses of isotope-labeled peptides are time-consuming and costly. To eliminate the need for using standards or calibration curves, herein we present a coulometric mass spectrometric (CMS) approach for absolute protein quantitation, based on the electrochemical oxidation of a surrogate peptide combined with mass spectrometric measurement of the oxidation yield. To demonstrate the utility of this method, several proteins were analyzed such as model proteins β-casein, and apomyoglobin as well as circadian clock protein KaiB isolated from Escherichia coli. In our experiment, tyrosine-containing peptides were selected as surrogate peptides for quantitation, considering the oxidizable nature of tyrosine. Our data showed that the results for surrogate peptide quantity measured by our method and by traditional isotope dilution method are in excellent agreement, with the discrepancy of 0.3-3%, validating our CMS method for absolute quantitation. Furthermore, therapeutic monoclonal antibody (mAb) could be quantified by our method as well. Due to the high specificity and sensitivity of MS and no need to use isotope-labeled peptide standards, our CMS method would be of high value for the absolute proteomic quantification.

Original languageEnglish (US)
Pages (from-to)7877-7883
Number of pages7
JournalAnalytical Chemistry
Volume92
Issue number11
DOIs
StatePublished - Jun 2 2020

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

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