Unit

UNIT 23.4 Quantitative Protein Analysis Using Enzymatic [18O]Water Labeling

  1. Mary Joan Castillo1,
  2. Kristy J. Reynolds2,
  3. Alexander Gomes1,
  4. Catherine Fenselau2,
  5. Xudong Yao1

Published Online: 1 APR 2014

DOI: 10.1002/0471140864.ps2304s76

Current Protocols in Protein Science

Current Protocols in Protein Science

How to Cite

Castillo, M. J., Reynolds, K. J., Gomes, A., Fenselau, C. and Yao, X. 2014. Quantitative Protein Analysis Using Enzymatic [18O]Water Labeling. Current Protocols in Protein Science. 76:23.4:23.4.1–23.4.9.

Author Information

  1. 1

    Department of Chemistry, University of Connecticut, Storrs, Connecticut

  2. 2

    Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland

Publication History

  1. Published Online: 1 APR 2014

Abstract

This unit describes the stepwise procedure for differential oxygen isotope labeling of peptides and mass spectrometric quantification of relative protein levels in comparative proteomic experiments. The [18O] labeling of peptides happens at the peptide C-terminus and is achieved via the enzymatic oxygen exchange of tryptic peptides via catalysis of immobilized trypsin. Experimental considerations in effective incorporation and stabilization of the oxygen labels are discussed. Methods for mass spectrometric quantification of peptides with differential [16O] and [18O] isotopes are presented. Curr. Protoc. Protein Sci. 76:23.4.1-23.4.9. © 2014 by John Wiley & Sons, Inc.

Keywords:

  • 18O-labeling;
  • enzymatic oxygen labeling;
  • quantitative proteomics;
  • immobilized trypsin;
  • back-exchange