J Biol Chem 2002 Apr 18;8(6) [epub ahead of print]
Peptides identify the critical hotspots involved in the biological activation of the insulin receptor.
Pillutla RC, Hsiao Kc K, Beasley JR, Brandt J, Ostergaard S, Hansen PH, Spetzler JC, Danielsen GM, Andersen AS, Brissette RE, Lennick M, Fletcher PW, Blume AJ, Schaffer L, Goldstein NI.
Molecular Biology, DGI BioTechnologies Inc, Edison, NJ 07040.
We used phage display to generate Surrogate peptides that define the hotspots involved in the protein:protein interaction between insulin and the insulin receptor. All of the peptides competed for insulin binding and had affinity constants in the high nanomolar to low micromolar range. Based on competition studies, peptides were grouped into non-overlapping Sites 1, 2 or 3. Some Site 1 peptides were able to activate the tyrosine kinase activity of the insulin receptor and act as agonists in the insulin-dependent fat cell assay, suggesting that Site 1 marks the hotspot involved in insulin-induced activation of the insulin receptor. On the other hand, Site 2 and 3 peptides were found to act as antagonists in the phosphorylation and fat cell assays. These data show that peptide display can be used to define the molecular architecture of a receptor and identify the critical regions required for biological activity in a site directed manner. |
