Identification of GIYWHHY as a novel peptide substrate for human p60(c-src) protein tyrosine kinase

We have recently determined that -Ile-Tyr- were the two critical residues as a peptide substrate for p60(c-src) protein tyrosine kinase (Leu, Q. et al., Lea Peptide Sci., 1995, 2, 289). Here, we report on the design and synthesis of a secondary 'one-bead, one-compound' combinatorial peptide library based on this dipeptide motif (XIYXXXX, where X = all 19 eukaryotic amino acids except for cysteine). This secondary library was screened for its ability to be phosphorylated by p60(c-src) PTK using [gamma(32)P]ATP as a tracer. Five of the strongest [P-32]-labeled peptide-beads were identified and microsequenced: GIYWHHY, KIYDDYE, EIYEENG, EIYEEYE, and YIYEEED. A solid-phase phosphorylation assay was used to evaluate the structure-activity relationship of GIYWHHY. It was determined that Ile(2), Ty(3), His(5), and His(6) were crucial for its activity as a substrate. Copyright (C) 1996 Elsevier Science Ltd.