Format

Send to

Choose Destination
J Proteome Res. 2014 Dec 5;13(12):5461-70. doi: 10.1021/pr500845u. Epub 2014 Sep 30.

Proteome-wide discovery of unknown ATP-binding proteins and kinase inhibitor target proteins using an ATP probe.

Author information

1
Laboratory of Proteome Research, National Institute of Biomedical Innovation , Ibaraki, Osaka 567-0085, Japan.

Abstract

ATP-binding proteins, including protein kinases, play essential roles in many biological and pathological processes and thus these proteins are attractive as drug targets. Acyl-ATP probes have been developed as efficient probes for kinase enrichment, and these probes have also been used to enrich other ATP-binding proteins. However, a robust method to identify ATP-binding proteins with systematic elimination of nonspecific binding proteins has yet to be established. Here, we describe an ATP competition assay that permitted establishment of a rigorous ATP-binding protein list with virtual elimination of nonspecific proteins. A total of 539 ATP-binding protein candidates were identified, including 178 novel candidates. In informatics analysis, ribosomal proteins were overrepresented in the list of novel candidates. We also found multiple ATP-competitive sites for several kinases, including epidermal growth factor receptor, serine/threonine-protein kinase PRP4 homologue, cyclin-dependent kinase 12, eukaryotic elongation factor 2 kinase, ribosomal protein S6 kinase alpha-1, and SRSF protein kinase 1. Using our cataloged ATP-binding protein list, a selectivity profiling method that covers the kinome and ATPome was established to identify off-target binding sites of ATP-competitive kinase inhibitors, staurosporine and crizotinib.

KEYWORDS:

ATP-binding proteins; ATP-competitive kinase inhibitor; Chemical proteomics

PMID:
25230287
DOI:
10.1021/pr500845u
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for American Chemical Society
Loading ...
Support Center