FIGURE 3.

Tryptic phosphopeptide maps of wild-type (WT) and mutant (knock-out; KO) forms of EphA2. Wild-type and mutant forms of EphA2 were expressed in EphA2-null endothelial cells via LZRS retroviral transduction. EphA2 receptors were immunoprecipitated, phosphorylated in the presence of [γ-³²P] ATP, and resolved by SDS-PAGE. Radioactive bands representing phosphorylated forms of EphA2 were excised, digested with trypsin, and subjected to two-dimensional chromatography. Four peptides were detected (labeled a-d) in both wild-type and EphA2-null reconstituted with wild-type EphA2 in Experiment 1. An additional phosphopeptide (labeled e) was identified in Experiment 2 when the first dimension chromatography was performed in the reverse direction. Peptides containing identified phosphorylated tyrosines are indicated on the left in two schematic representations, one for each set of experiments.