Research
The Insulin Receptor Kinase Activation loop
Using the holoreceptor from human placenta and the soluble kinase expressed in Sf9 insect cells (baculovirus system), we are characterizing the dual specificity of the insulin receptor and its effects on signal transduction. With the help of HPLC phosphopeptide mapping, we are able to analyze specific phosphorylation sites in the insulin receptor and its substrates. By site-directed mutagenesis, we created a number of mutated receptor kinases to determine the influence of single amino acid residues. Recombinant fragments of IR domains and in vivo substrates (IRS-1, Gab-1) help to understand how the receptor gets activated and interacts with downstream signaling molecules. We have cloned and expressed a glutathione S-transferase (GST)-dimerized insulin receptor kinase as a novel model for the holoreceptor.
In addition to the insulin receptor, we are studying soluble tyrosine kinases of the Insulin Like Growth Factor-1 Receptor (IGF-1R), the Epidermal Growth Factor Receptor (EGFR) and the BDNF / NT-3 Growth Factors Receptor (TrkB). All of these show a dual specificity, leading to the hypothesis that serine phosphorylation of receptor tyrosine kinases might be a general phenomenon. The IGF-1R kinase has been dimerized by GST in the same way as the insulin receptor.
Interested?
Our focus of interest is the catalytic mechanism of receptor tyrosine kinases. One main object of investigation is the Insulin Receptor (IR). Non insulin dependent diabetes mellitus (NIDDM) is characterized by a poor or missing signal transduction, resulting in an impaired cellular glucose uptake. This might occur on the stage of the insulin receptor or at a post-receptor level. Serine phosphorylation of the receptor or receptor substrates is thought to inhibit the insulin signaling in the cell. Recently, we have identified two serine residues which are autophosphorylated by the receptor itself. Y1151 in the activation loop plays a key role in this serine kinase activity. Moreover, a recombinant fragment of IRS-1 is also phosphorylated on serine and threonine residues by the insulin receptor kinase. Therefore, the serine kinase activity of the insulin receptor could play an important role of down-regulating the insulin signal.
