Peptide microarrays for the detection of molecular interactions in cellular signal transduction.
The formation of protein complexes is a hallmark of cellular signal transduction. Here, we show that peptide microarrays provide a robust and quantitative means to detect signalling-dependent changes of molecular interactions. Recruitment of a protein into a complex upon stimulation of a cell leads to the masking of an otherwise exposed binding site. In cell lysates this masking can be detected by reduced binding to a microarray carrying a peptide that corresponds to the binding motif of the respective interaction domain. The method is exemplified for the lymphocyte-specific tyrosine kinase 70 kDa zeta-associated protein binding to a bis-phosphotyrosine-motif of the activated T-cell receptor via its tandem SH2 domain. Compared to established techniques, the method provides a significant shortcut to the detection of molecular interactions.
Stoevesandt
Oda
O
Institute for Cell Biology, University of Tübingen, Germany.
Elbs
Martin
M
Köhler
Karsten
K
Lellouch
Annemarie C
AC
Fischer
Rainer
R
André
Thomas
T
Brock
Roland
R
D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D006825:Hybridomas; D008213:Lymphocyte Activation; D051379:Mice; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D040081:Protein Array Analysis; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence; D013601:T-Lymphocytes / Q000378:metabolism; D051746:ZAP-70 Protein-Tyrosine Kinase; D018909:src Homology Domains
D005456:Fluorescent Dyes; D010455:Peptides; D011948:Receptors, Antigen, T-Cell; D011505:Protein-Tyrosine Kinases; D051746:ZAP-70 Protein-Tyrosine Kinase; C497262:Zap70 protein, mouse