The mapping of the Lyn kinase binding site of the common beta subunit of IL-3/granulocyte-macrophage colony-stimulating factor/IL-5 receptor.
It has been shown that a membrane-proximal region within common beta (betac) receptor of IL-3/granulocyte-macrophage CSF/IL-5 (amino acids 450-517) is important for Lyn binding. We have shown previously that Lyn kinase is physically associated with the IL-5R betac subunit in unstimulated cells. The result suggests that this association involves binding modules that are not activation or phosphorylation dependent. The objective of this study was to map the exact Lyn binding site on betac. Using overlapping and/or sequential peptides derived from betac 450-517, we narrowed down the Lyn binding site to nine amino acid residues, betac 457-465. The P-->A mutation in this region abrogated the binding to Lyn, indicating a critical role of proline residues. We created a cell-permeable Lyn-binding peptide by N-stearation. This cell-permeable peptide blocked the association of Lyn, but not Jak2 with betac in situ. We also investigated the betac binding site of Lyn kinase. Our results suggest that the N-terminal unique domain of Lyn kinase is important for binding to betac receptor. To our knowledge, this is the first molecular identification of the Lyn binding site of betac receptor. This finding may help develop specific inhibitors of Lyn-coupled signaling pathways.
Adachi
T
T
Department of Internal Medicine, Division of Allergy and Immunology, University of Texas Medical Branch, Galveston, TX 77555, USA.
Pazdrak
K
K
Stafford
S
S
Alam
R
R
D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D053648:Cytokine Receptor Common beta Subunit; D006801:Humans; D066298:In Vitro Techniques; D007962:Leukocytes / Q000276:immunology / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010449:Peptide Mapping; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018123:Receptors, Interleukin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016185:Receptors, Interleukin-3 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D053649:Receptors, Interleukin-5; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D019061:src-Family Kinases / Q000378:metabolism*