HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: M600105-MCP200v1 5/9/1610    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Glossary Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Wu, S.-L. Articles by Karger, B. L. Search for Related Content PubMed PubMed Citation Articles by Wu, S.-L. Articles by Karger, B. L. Social Bookmarking                      What's this?

Molecular & Cellular Proteomics 5:1610-1627, 2006.
© 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Research

Dynamic Profiling of the Post-translational Modifications and Interaction Partners of Epidermal Growth Factor Receptor Signaling after Stimulation by Epidermal Growth Factor Using Extended Range Proteomic Analysis (ERPA)^*,S

Shiaw-Lin Wu^,, Jeongkwon Kim^,, Russell W. Bandle^¶, Lance Liotta^||, Emanuel Petricoin^|| and Barry L. Karger^,**

From the Barnett Institute, Northeastern University, Boston, Massachusetts 01225, ^¶ Laboratory of Pathology, NCI, National Institutes of Health, Bethesda, Maryland 20892, and ^|| Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, Virginia 20110

In a recent report, we introduced Extended Range Proteomic Analysis (ERPA), an intermediate approach between top-down and bottom-up proteomics, for the comprehensive characterization at the trace level (fmol level) of large and complex proteins. In this study, we extended ERPA to determine quantitatively the temporal changes that occur in the tyrosine kinase receptor, epidermal growth factor receptor (EGFR), upon stimulation. Specifically A 431 cells were stimulated with epidermal growth factor after which EGFR was immunoprecipitated at stimulation times of 0, 0.5, 2, and 10 min as well as 4 h. High sequence coverage was obtained (96%), and methods were developed for label-free quantitation of phosphorylation and glycosylation. A total of 13 phosphorylation sites were identified, and the estimated stoichiometry was determined over the stimulation time points, including Thr(P) and Ser(P) sites in addition to Tyr(P) sites. A total of 10 extracellular domain N-glycan sites were also identified, and major glycoforms at each site were quantitated. No change in the extent of glycosylation with stimulation was observed as expected. Finally potential binding partners to EGFR were identified based on changes in the amount of protein pulled down with EGFR as a function of time of stimulation. Many of the 19 proteins identified are known binding partners of EGFR. This work demonstrates that comprehensive characterization provides a powerful tool to aid in the study of important therapeutic targets. The detailed molecular information will prove useful in future studies in tissue.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				N. A. Watkins, A. Gusnanto, B. de Bono, S. De, D. Miranda-Saavedra, D. L. Hardie, W. G. J. Angenent, A. P. Attwood, P. D. Ellis, W. Erber, et al. A HaemAtlas: characterizing gene expression in differentiated human blood cells Blood, May 7, 2009; 113(19): e1 - e9. [Abstract] [Full Text] [PDF]

				M. Mann and N. L. Kelleher Mass Spectrometry Special Feature: Precision proteomics: The case for high resolution and high mass accuracy PNAS, November 25, 2008; 105(47): 18132 - 18138. [Abstract] [Full Text] [PDF]

				A. J. VanMeter, A. S. Rodriguez, E. D. Bowman, J. Jen, C. C. Harris, J. Deng, V. S. Calvert, A. Silvestri, C. Fredolini, V. Chandhoke, et al. Laser Capture Microdissection and Protein Microarray Analysis of Human Non-small Cell Lung Cancer: Differential Epidermal Growth Factor Receptor (EGPR) Phosphorylation Events Associated with Mutated EGFR Compared with Wild Type Mol. Cell. Proteomics, October 1, 2008; 7(10): 1902 - 1924. [Abstract] [Full Text] [PDF]

				J. J. Lammerts van Bueren, W. K. Bleeker, A. Brannstrom, A. von Euler, M. Jansson, M. Peipp, T. Schneider-Merck, T. Valerius, J. G. J. van de Winkel, and P. W. H. I. Parren The antibody zalutumumab inhibits epidermal growth factor receptor signaling by limiting intra- and intermolecular flexibility PNAS, April 22, 2008; 105(16): 6109 - 6114. [Abstract] [Full Text] [PDF]

				D. Pflieger, M. A. Junger, M. Muller, O. Rinner, H. Lee, P. M. Gehrig, M. Gstaiger, and R. Aebersold Quantitative Proteomic Analysis of Protein Complexes: Concurrent Identification of Interactors and Their State of Phosphorylation Mol. Cell. Proteomics, February 1, 2008; 7(2): 326 - 346. [Abstract] [Full Text] [PDF]

				D. D. Vadysirisack, E. S.-W. Chen, Z. Zhang, M.-D. Tsai, G.-D. Chang, and S. M. Jhiang Identification of in Vivo Phosphorylation Sites and Their Functional Significance in the Sodium Iodide Symporter J. Biol. Chem., December 21, 2007; 282(51): 36820 - 36828. [Abstract] [Full Text] [PDF]

				T. Stasyk, N. Schiefermeier, S. Skvortsov, H. Zwierzina, J. Peranen, G. K. Bonn, and L. A. Huber Identification of Endosomal Epidermal Growth Factor Receptor Signaling Targets by Functional Organelle Proteomics Mol. Cell. Proteomics, May 1, 2007; 6(5): 908 - 922. [Abstract] [Full Text] [PDF]