THE 3-DIMENSIONAL SOLUTION STRUCTURE OF THE SH2 DOMAIN FROM P55(BLK) KINASE

Signal transduction in B cells is mediated, in part, by the interactio n of the cytoplasmic components of the antigen receptor complex and va rious members of the src family tyrosine kinases. Key to this process appears to be the interaction of the tyrosine kinase SH2 domains with the tyrosine-phosphorylated cytoplasmic domain of Ig-alpha, a disulfid e-bonded heterodimeric (with Ig-beta or Ig-gamma) transmembrane protei n that noncovalently associates with the antigen receptor immunoglobin chains. In addition to binding to the phosphorylated cytoplasmic doma ins of Ig-alpha and Ig-beta, blk and fyn(T), two members of the si-e f amily kinases, have been shown to bind overlapping but distinct sets o f phosphoproteins [Malek & Desiderio (1993) J. Biol. Chem. 268, 22557- 22565]. A comparison of their three-dimensional structures may elucida te the apparently subtle differences required for phosphoprotein discr imination. To begin characterizing the blk/fyn/phosphosphoprotein inte ractions, we have determined the three-dimensional solution structure of the SH2 domain of blk kinase by nuclear magnetic resonance (NMR) sp ectroscopy. H-1, C-13, and N-15 resonances of the SH2 domain of blk ki nase were assigned by analysis of multidimensional, double- and triple -resonance NMR experiments. Twenty structures of the blk SH2 domain we re refined with the program X-PLOR using a total of 2080 experimentall y derived conformational restraints. The structures converged to a roo t-mean-squared (rms) distance deviation of 0.51 and 0.95 Angstrom for the backbone atoms and for the non-hydrogen atoms, respectively. The b lk SH2 domain adopts the prototypical SH2 fold. Structurally, blk SH2 is most similar to the crystal structure of the v-src SH2 domain [Waks man et al. (1993) Nature 358, 646-653] and superimposes on the crystal structure with an rmsd of 1.52 Angstrom for the backbone atoms. The l argest deviations occur in the four loops interconnecting beta-strands A-E, which are the least well-defined regions in the NMR structure. E xclusion of these loops lowers this rmsd to 0.82 Angstrom. The conform ation of the BC loop in the blk SH2 domain is similar to the open conf ormation in the apo lck SH2 domain, suggesting that, like the lck SH2 domain, the blk SH2 domain may have a gated phosphopeptide binding sit e. Finally, it is proposed that the amino acid substitution of Lys 88 (blk) for Glu [fyn(T)] is important for the observed differences in sp ecificity between blk and fyn(T) SH2 domains.