Activity profiling of platelets by chemical proteomics

Chemical proteomics or activity based proteomics is a functional proteomics technology where molecular probes are used to target a selective group of functionally related proteins. Its emergence has enabled specific targeting of subproteomes, overcoming the limitations in dynamic range of traditiona...

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Bibliographic Details
Published in:Proteomics (Weinheim) 2009, Vol.9 (1), p.40-50
Main Authors: Wong, Jason W.H, McRedmond, James P, Cagney, Gerard
Format: Article
Language:English
Subjects:
Adenosine Triphosphate - metabolism
Affinity chromatography
Blood Platelets - chemistry
Blood Platelets - metabolism
Chemical proteomics
Chromatography, Affinity
Cyclic AMP - metabolism
Cyclic GMP - metabolism
Humans
Label‐free quantification
Nucleotides
Nucleotides - metabolism
Platelet Activation
Platelets
Protein Binding
Proteome - analysis
Proteome - metabolism
Proteomics - methods
Substrate Specificity
Tandem Mass Spectrometry
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Summary:Chemical proteomics or activity based proteomics is a functional proteomics technology where molecular probes are used to target a selective group of functionally related proteins. Its emergence has enabled specific targeting of subproteomes, overcoming the limitations in dynamic range of traditional large-scale proteomics experiments. Using a chemical proteomics strategy, we attempt to differentially profile the nucleotide-binding proteome of active and resting platelets. We apply an affinity chromatography protocol using immobilized adenosine triphosphate, cyclic adenosine monophosphate, and cyclic guanosine monophosphate. The specificity of the immobilized nucleotides was demonstrated by competitive assays and by immunoblotting. LC coupled MS/MS was applied to identify the proteins recovered by our chemical proteomics strategy. When compared to a standard set of platelet lysate proteins, we confirmed that enrichment for nucleotide-binding proteins was indeed taking place. Finally, by employing label-free MS-based comparative quantification, we found a small number of platelet proteins that show statistically significant difference between the active and resting nucleotide-binding proteome.
ISSN:1615-9853
1615-9861
DOI:10.1002/pmic.200800185