Identification of GIYWHHY as a novel peptide substrate for human p60c-src protein tyrosine kinase.
We have recently determined that -Ile-Tyr- were the two critical residues as a peptide substrate for p60c-src protein tyrosine kinase (Lou, Q. et al., Lett. 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 p60c-src PTK using [gamma 32P]ATP as a tracer. Five of the strongest [32P]-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 Ile2, Tyr3, His5, and His6 were crucial for its activity as a substrate.
Lou
Q
Q
Arizona Cancer Center, Tucson, USA.
Leftwich
M E
ME
Lam
K S
KS
D000255:Adenosine Triphosphate / Q000032:analysis; D000595:Amino Acid Sequence; D001345:Autoradiography; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D017421:Sequence Analysis; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D010455:Peptides; D010761:Phosphorus Radioisotopes; D000255:Adenosine Triphosphate; D011505:Protein-Tyrosine Kinases; D016392:Proto-Oncogene Proteins pp60(c-src)