SAP requires a threonine residue at position −2 in the consensus motif T(I/V)pYxx(V/I) to stabilize both phosphotyrosine- and non–phosphotyrosine-dependent binding to CD150.

(A) COS-7 cells transiently transfected with CD150 or CD150 with the indicated threonine-alanine substitutions plus SAP-FLAG with or without Fyn coexpression. CD150 was immunoprecipitated followed by Western blotting with the indicated antibodies. (B) Yeast 2-hybrid analysis of SAP binding to non–tyrosine-phosphorylated CD150 T-A constructs. ▪, +SAP; ■, +pBridge. Quantitation of β-galactosidase activity was performed as described in “Materials and methods.”