Please cite as: CSH Protocols; 2008; doi:10.1101/pdb.prot4610

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Alkaline Phosphatase Treatment of Phosphopeptides: In-Solution Dephosphorylation prior to MALDI-MS Analysis

Hanno Steen, Allan Stensballe, and Ole N. Jensen

This protocol was adapted from "Proteomic Methods for Phosphorylation Site Mapping," Chapter 9, in Proteins and Proteomics (ed. Simpson). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2003.


INTRODUCTION

The use of the enzyme alkaline phosphatase allows identification of phosphopeptides in a mixture of predominantly nonphosphopeptides. Using a MALDI-MS instrument, the masses of peptides are acquired both before and after alkaline phosphatase treatment, which removes phospho-moieties from serine, threonine, and/or tyrosine. (Any peptide whose mass decreases by 80 Da, or a multiple thereof, is a phosphopeptide.) An advantage of using MALDI-MS for these experiments is that the peptide ions produced tend to be singly charged rather than multiply charged (as with ESI), thus making the interpretation easier. This protocol describes a method for in-solution dephosphorylation prior to MALDI-MS analysis.


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This article has been cited by other articles:


Home page
 CSHL ProtocolsHome page
H. Steen, A. Stensballe, and O. N. Jensen
Alkaline Phosphatase Treatment of Phosphopeptides: In-Solution Dephosphorylation after MALDI-MS Analysis
CSH Protocols, May 1, 2008; 2008(6): pdb.prot4611 - pdb.prot4611.
[Abstract] [Full Text]

Home page
 CSHL ProtocolsHome page
H. Steen, A. Stensballe, and O. N. Jensen
Alkaline Phosphatase Treatment of Phosphopeptides: On-Probe Dephosphorylation after MALDI-MS Analysis
CSH Protocols, May 1, 2008; 2008(6): pdb.prot4612 - pdb.prot4612.
[Abstract] [Full Text]