Molecular cloning and expression of human Grap-2, a novel leukocyte-specific SH2- and SH3-containing adaptor-like protein that binds to Gab-1

The SH2- and SH3-containing adaptor molecules serve to recruit cytosolic signal-transducing molecules to the activated receptor tyrosine kinases. In this study, we report the molecular cloning of a novel adaptor-like protein, Grap-2 (Grb-2 related adaptor protein 2), using the multisubstrate docking protein Gab-1 as bait in the yeast two-hybrid system. Sequence analysis revealed that Grap-2 contains a SH3-SH2-SH3 structure that has a high degree of sequence homology to those of the Grb-2 and Grap adaptor molecules. However, unlike in Grap and Grb-2, the SH2 and the C-terminal SH3 domains of Grap-2 are separated by a 120-amino-acid glutamine-rich sequence that shows no apparent homology to any known molecule or structural motif. The C-terminal SH3 domain of Grap-2 alone is sufficient to bind to Gab-1. Furthermore, Northern blot analysis demonstrated that Grap-2 has two major transcripts of 1.4 and 4.0 kb that can only be detected in tissues rich in leukocytes and in two leukemia cell lines. This highly restricted pattern of expression suggests that Grap-2 may participate in leukocyte-specific protein tyrosine kinase signaling.