Identification of a binding sequence for the 14-3-3 protein within the cytoplasmic domain of the adhesion receptor, platelet glycoprotein Ib alpha.
The zeta-form 14-3-3 protein (14-3-3zeta) regulates protein kinases and interacts with several signaling molecules. We reported previously that a platelet adhesion receptor, glycoprotein (GP) Ib-IX, was associated with a 29-kDa protein with partial sequences identical to 14-3-3zeta. In this study, the interaction between GPIb-IX and recombinant 14-3-3zeta is reconstituted. Further, we show that the 14-3-3zeta binding site in GPIb is within a 15 residue sequence at the C terminus of GPIb-alpha, as indicated by antibody inhibition and direct binding of 14-3-3zeta to synthetic GPIb-alpha cytoplasmic domain peptides. The 14-3-3zeta binds to recombinant wild type GPIb-IX but not to the GPIb-alpha mutants lacking C-terminal 5 or more residues, suggesting that the C-terminal 5 residues of GPIb-alpha are critical. Similarity between the GPIb-alpha C-terminal sequence and the serine-rich regions of Raf and Bcr kinases suggests a possible serine-rich recognition motif for the 14-3-3 protein.
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Department of Vascular Biology, Scripps Research Institute, La Jolla, California 92037, USA.
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D048948:14-3-3 Proteins; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D001665:Binding Sites; D016466:CHO Cells; D002460:Cell Line; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D006224:Cricetinae; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D015723:Gene Library; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine; D015398:Signal Transduction; D014162:Transfection; D014407:Tumor Cells, Cultured; D014446:Tyrosine 3-Monooxygenase
D048948:14-3-3 Proteins; D000906:Antibodies; D017931:DNA Primers; D010455:Peptides; D019038:Platelet Glycoprotein GPIb-IX Complex; D011506:Proteins; D011994:Recombinant Proteins; D012694:Serine; D014446:Tyrosine 3-Monooxygenase