Identification of the critical features of a small peptide inhibitor of JNK activity.
The c-Jun N-terminal kinases (JNKs) are a subfamily of the mitogen-activated protein kinases (MAPKs). Although progress in evaluating the functions of other MAPKs has been facilitated by the characterization of specific inhibitors, no JNK-directed inhibitor is commercially available. We have identified a 21-amino acid peptide inhibitor of activated JNKs, based on amino acids 143-163 of the JNK-binding domain (JBD) of the JNK scaffolding protein, JNK-interacting protein-1 (JIP-1). This peptide, I-JIP (Inhibitor of JNK-based on JIP-1), inhibited JNK activity in vitro toward recombinant c-Jun, Elk, and ATF2 up to 90%. A truncated I-JIP (TI-JIP), the C-terminal 11 amino acids of I-JIP, directly interacted with recombinant JNKs but not its substrates as shown by surface plasmon resonance analysis. Scanning alanine replacement within truncated I-JIP identified 4 residues (Arg-156, Pro-157, Leu-160, or Leu-162) as independently critical for inhibition. JBD peptide sequences from JIP-2 and JIP-3 shared these critical residues and accordingly were effective JNK inhibitors. In contrast, peptides based on the JBDs of ATF2 and c-Jun inhibited JNK activity by <40%, which agreed with their lack of homology to the critical Arg-156 and Pro-157. These studies thus define a small peptide inhibitor sequence of JNKs based on the JIP proteins.
Barr
Renae K
RK
Department of Biochemistry, University of Western Australia, Crawley 6009, Australia.
Kendrick
Tulene S
TS
Bogoyevitch
Marie A
MA
D051698:Activating Transcription Factor 2; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases; D007700:Kinetics; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D020349:Surface Plasmon Resonance; D014157:Transcription Factors / Q000378:metabolism; D050807:ets-Domain Protein Elk-1; D048051:p38 Mitogen-Activated Protein Kinases
D051698:Activating Transcription Factor 2; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D010455:Peptides; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases