Tyrosine Kinase Peptide 1

Definition
Peptide sequence KVEKIGEGTYGVVYK is derived from the amino acid residues CDC26-20. It is considered to be a generic substrate for various tyrosine kinase peptide (TPK).

Discovery
Lou, Q. et al., in 1995 determined the two amino acids Ile and Tyr were the two critical residues as a peptide substrate for p60c-src protein tyrosine kinase 1. Subsequently in 1996 they reported 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) 2. Normanno in 2003 analysed epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) 3.

Structural Characteristics
Sequence of TPK 1 is, 5 - TMR - Lys - Val - Glu - Lys - Ile - Gly - Glu - Gly - Thr - Tyr - Gly - Val - Val - Tyr - Lys - OH. Lou Q. et al., in 1996 screened a secondary library for TPK 1for its ability to be phosphorylated by p60c-src PTK using [?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 2. Using various combinatorial peptide library approaches, Wu et al., 1997 and Schmitz et al., 1996 were successful in identifying some relatively potent peptide substrates for a number of PTKs. For example, YIYGSFK, GIYWHHY, and EEIYGEFF for Src PTK have Kms ranging between 25-55 M 4,5.

Mode of Action
PTKs are key enzymes in many signal transduction pathways and play a crucial role in many cell regulatory processes. It is estimated that there are over 2000 protein serine/threonine kinases and close to 100 different PTKs encoded by the human genome. Numerous small molecule inhibitors for various PTKs have been developed. PTKs play a crucial role in many cell regulatory processes such as mitogenesis, differentiation and development, oncogenesis, angiogenesis, cell survival and apoptosis, cell shape and adhesion, cell cycle control, growth control, T-cell and B-cell activation, responses to extracellular stimuli, neurotransmitter signaling, platelet activation, transcription control, and glucose uptake 4. Synthetic peptides derived from autophosphorylation sites or phosphorylation sites of protein substrates are often poor substrates for PTK. They can serve as important reagents to study the mechanism of action of the enzyme, to map the pharmacophores of the substrate binding pocket, and to be used as leads for the design of peptidomimetic inhibitors 6. The native peptide derived from pp60src, and considered to be a generic substrate for various protein tyrosine kinases. The fluorescent and biotinylated peptides are used to design a variety of assays for PTKs.

The epidermal growth factor receptor (EGFR) has been identified as a relevant target as it is involved in regulating several cellular functions important in the proliferation and survival of cancer cells, is commonly expressed at high levels in a range of tumors, and high expression is often related to poor prognosis. EGFR is a member of the ErbB family of receptors which also includes ErbB-2, ErbB-3, and ErbB-4. These receptors form dimers of the same type (homodimers) or with other family members (heterodimers), each combination resulting in different downstream effects. Some of the most advanced targeted agents in development are the EGFR tyrosine kinase inhibitors (EGFR-TKIs), of which ZD1839 ('Iressa') is an example. Tyrosine kinase inhibitors affect EGFR signaling and may also affect signaling of other ErbB family members 3.

Functions
Cell signaling, PTKs (as well as serine and threonine kinases) are crucial enzymes in many cell signalling pathways and are altered or overexpressed in many disease states. Inhibitors against some of these enzymes may be useful for the treatment of a number of diseases.

Assays for PTKs, the fluorescent and biotinylated peptides are used to design a variety of assays for PTKs 2.

Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs): EGFR-TKIs have been shown in preclinical studies to increase the efficacy of cytotoxic drugs and Phase III trials of such combinations are ongoing. On the basis that different signal transduction pathways contribute to the control of tumor growth, future therapeutic approaches are likely to involve combination of different targeted agents 3.

References
1. Lou Q, Leftwich ME, Lam KS (1996). Identification of GIYWHHY as a novel peptide substrate for human p60c-src protein tyrosine kinase. Bioorg Med Chem., 4(5):677-682.
2. Lou Q, Wu J, Salmon SE, Lam KS (1995). Structure-activity relationship of a novel peptide substrate for p60c-src protein tyrosine kinase. Lett. Custom Peptide Sci., 2:289:296.
3. Normanno N, Maiello MR, De Luca A (2003). Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs): simple drugs with a complex mechanism of action. J Cell Physiol., 194(1):13-9.
4. H unter T (1997). Oncoprotein networks. Cell, 88:333-346.
5. Wu JJ, Phan H and Lam KS (1998). Comparison of the intrinsic kinase activity and substrate specificity of c-Abl and Bcr-Abl. Bioorg. Med. Chem. Lett., 8: 2279-2284.
6. Schmitz R, Baumann G and Gram H (1996). Catalytic specificity of phosphotyrosine kinases Blk, Lyn, c-Src and Syk as assessed by phage display. J. Mol. Biol., 260(5):664-677.

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