Identification of IGF-II-Binding Site on the Quaternary 3-D Structure of the Insulin Receptor
Abstract
Gold-labeled IGE-lI has been prepared in micro-scale by reacting IGE-lI with sulfo-NHS- Nanogold, and purified by size-exclusion HPLC. Two gold-labeled TOE-II products, differing in electrophoretic mobility under acidic condition, were obtained. Soth products were derivatized with the Nanogold at their N-terminal alanine. Insulin receptor proteins, IR-A and IR-B have been extracted and purified from transfectant cells. As expected from previous studies in the transfectant cells, IR-A in contrast to IR-B was activated by IcE-II comparable to that by insulin. The two gold-labeled TOE-lI derivatives when tested in a preliminary study for their activity on IR-A were found to have a good level of activity in stimulating the autophosphorylation of IR-A. In summary, methods have been established to prepare and purify the reagents, namely the gold-labeled TOE-IT and the insulin receptor proteins, that are needed to form the ligand-receptor complex for the identification of the IGE-IT-binding site of the insulin receptor by STEM reconstruction of the 3-D guaternary structure of the complex.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 01, 2001
- Accession Number
- ADA392396
Entities
People
- Cecil C. Yip
Organizations
- University of Toronto