In vitro characterization of major ligands for Src homology 2 domains derived from protein tyrosine kinases, from the adaptor protein SHC and from GTPase‐activating protein in Ramos B cells

Antigen receptors of B lymphocytes transmit their activation signal to the cell interior by associating with and activation of specific non‐receptor tyrosine kinases. Most of these kinases as well as other cytoplasmic effectors contain at least one Src homology 2 (SH2) domain, known to bind tyrosine‐phosphorylated proteins. We examined the binding specificity of SH2 domains from different signaling molecules in B cells and found that each of the SH2 domains tested bound distinct subsets of stimulation‐dependent phosphoproteins in vitro. SH2 domains from Src‐like tyrosine kinases bound predominantly to the HS1 phosphoprotein. The tandem SH2 domains of the ZAP‐70 tyrosine kinase bound to phosphorylated Ig‐β but only weakly to Ig‐α. Also the SHC‐derived SH2 domain formed complexes with the tyrosine‐phosphorylated Ig‐α/β heterodimer, while the C‐ and N‐terminal SH2 domains of GTPase‐activating protein displayed completely different binding preferences. These results suggest that cytoplasmic effector molecules can be recruited to the activated B cell receptor in an SH2‐phosphotyrosine‐mediated manner. The data also provide a possible explanation for the notion that Ig‐α and Ig‐β might couple to different biochemical pathways.