LAUSR

Proteolytic cleavage of membrane proteins can alter their functions depending on the cleavage sites. We recently demonstrated that membrane type 1 matrix metalloproteinase (MT1-MMP ) converts the tumor suppressor EphA2 into an oncogenic signal transducer through EphA2 cleavage. The cleaved EphA2 fragment that remains at the cell surface may be a better target for cancer therapy than intact EphA2. To analyze the cleavage site(s) of EphA2, we purified the fragments from tumor cells expressing MT1-MMP and Myc- and 6× His-tagged EphA2 by two-step affinity purification . The purified fragment was digested with trypsin to generate proteolytic peptides , and the amino acid sequences of these peptides were determined by nano-LC-mass spectrometry to identify the MT1-MMP-mediated cleavage site(s) of EphA2.