The vasculotropic pathogenic bacteria Bartonella henselae secretes 7 Bep effector proteins (BepA-G) and injects these proteins into host cells using a type Ⅳ secretion system. Among these effector proteins, BepD-F contain multiple copies of Glu-Pro-Ile-Tyr-Ala (EPIYA) motifs in their N-terminal regions. After injection into host cells, the tyrosine residues in these EPIYA motifs will be phosphorylated by SFK, and the tyrosine-phosphorylated EPIYA motifs can subsequently interact with various SH2 domain-containing proteins from host cell and interfere with the SH2 signaling pathways of the host cells. In addition to B. henselae Bep effector proteins, numerous pathogenic bacteria effector proteins had been identified previously to contain EPIYA motifs. They were also found to utilize their EPIYA motifs to disrupt the SH2-signaling pathways of respective host cells, although they share no obvious overall sequence similarities. In contrast, only 6 proteins containing EPIYA motifs were found in human proteome, much lower than random occurrences, probably due to its toxicity in human cells. JAM-A is one of EPIYA motif containing proteins occurring in blood platelet, which can recruit Csk to avoid thrombus formation. BepE was reported to interact with Csk and disturb Csk-related signaling pathways. Here we reported the crystal structures of complexes between Csk and the phosphorylated EPIYA motifs from BepE and JAM-A, and biophysical measurements of their binding affinities. The complex structures show that Csk binds with two phosphopeptides in a similar way. The Csk SH2 domain binds with phosphotyrosine. Phosphopeptides are perpendicular to β-sheets in SH2 domain. Results of SPR show that the affinity of BepE with Csk is much higher than that of JAM-A. These results suggest that the EPIYA motif of toxic proteins binds with the SH2 domain of human Csk by a higher affinity, then disturb the SH2 domain proteins involved signaling pathways. The results reported here laid a solid foundation for our understanding of the structural basis of how toxic EPIYA motifs interrupt the SH2 signaling pathways of host cells.