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J Proteome Res. 2006 Aug;5(8):2019-24.

Selective ion tracing and MSn analysis of peptide digests from FSBA-treated kinases for the analysis of protein ATP-binding sites.

Author information

1
Proteomics & Mass Spectrometry Laboratory, ISPAAM, National Research Council, 80147 Naples, Italy.

Abstract

Kinases play a key role in many cellular processes by catalyzing the transfer of phosphoryl groups from ATP to a broad number of substrates, including amino acids on target proteins. The reagent 5'-fluorosulfonylbenzoyl-5'-adenosine (FSBA) has been widely used to identify ATP-binding sites in kinases since it reacts with nucleophilic amino acids occurring within these motifs, determining a mass increase of 433 Da. In this study, we present a versatile MS approach that has been developed to recognize labeled peptides generated after enzymatic digestion of FSBA-treated kinases. Using selective ion tracing and MS(2)/MS(3) experiments, we were able to easily identify peptides occurring at protein ATP-binding sites, also affording a complete characterization of the modified amino acids. This methodology may be used in the development of future parent ion scanning-based applications directed to large scale analysis of kinases within complex protein mixtures.

PMID:
16889425
DOI:
10.1021/pr0601234
[Indexed for MEDLINE]

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